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Aop: 272

AOP Title

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Direct deposition of ionizing energy onto DNA leading to lung cancer

Short name:

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Ionizing energy leading to lung cancer

Graphical Representation

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Authors

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Samantha Sherman1, Zakara Said1, Baki Sadi1, Carole Yauk1,2, Danielle Beaton3, Ruth Wilkins1 Robert Stainforth1, Vinita Chauhan1,*

Consumer and Clinical Radiation Protection Bureau, Health Canada, Ottawa, ON, Canada

2 Department of Biology, Carleton University, Ottawa, ON, Canada

Canadian Nuclear Laboratories, Chalk River, ON, Canada

*Corresponding author: Vinita Chauhan (vinita.chauhan@canada.ca)

 

 

 

Point of Contact

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Vinita Chauhan   (email point of contact)

Contributors

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  • Vinita Chauhan

Status

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Author status OECD status OECD project SAAOP status
Under development: Not open for comment. Do not cite


This AOP was last modified on September 06, 2019 15:15

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Revision dates for related pages

Page Revision Date/Time
Direct Deposition of Energy September 06, 2019 08:56
Increase, DNA strand breaks September 02, 2019 20:04
N/A, Inadequate DNA repair September 02, 2019 20:19
Increase, Mutations September 02, 2019 20:25
Increase, Chromosomal aberrations August 26, 2019 11:20
Increase, Cell Proliferation September 05, 2019 09:27
Increase, lung cancer September 02, 2019 21:06
Energy Deposition leads to Increase, DNA strand breaks September 04, 2019 13:51
Energy Deposition leads to Increase, Mutations September 04, 2019 14:03
Energy Deposition leads to Increase, Chromosomal aberrations September 04, 2019 14:07
Increase, DNA strand breaks leads to N/A, Inadequate DNA repair September 04, 2019 14:08
Energy Deposition leads to Increase, lung cancer September 04, 2019 15:10
N/A, Inadequate DNA repair leads to Increase, Mutations September 11, 2019 16:37
Increase, DNA strand breaks leads to Increase, Mutations September 05, 2019 18:58
Increase, DNA strand breaks leads to Increase, Chromosomal aberrations July 03, 2019 15:16
Increase, Mutations leads to Increase, lung cancer September 04, 2019 15:09
N/A, Inadequate DNA repair leads to Increase, Chromosomal aberrations September 04, 2019 15:06
Increase, Mutations leads to Increase, Cell Proliferation September 04, 2019 15:07
Increase, Chromosomal aberrations leads to Increase, lung cancer September 04, 2019 15:09
Increase, Chromosomal aberrations leads to Increase, Cell Proliferation September 04, 2019 15:08
Increase, Cell Proliferation leads to Increase, lung cancer September 04, 2019 15:08
Ionizing Radiation May 07, 2019 12:12

Abstract

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Despite its widespread recognition in chemical toxicology, the adverse outcome pathway (AOP) framework has not been fully explored in the radiation field to guide relevant research and subsequent risk assessment.  Development of a radiation relevant AOP is described here using a case example of lung cancer.  Lung cancer is a major public health problem world-wide, causing the deaths of an estimated 1.5 million people annually; it imposes a major health-care burden. Numerous environmental factors are known contributors including both chemical (eg. asbestos, air pollution and arsenic) and radiation stressors (eg. radon  gas).  Radon gas is the second leading cause of lung cancer in North America. Evidence suggests that environmental and indoor radon exposure constitutes a significant public health problem. The mechanism of lung cancer development from exposure to radon gas is unclear. Data suggest that cytogenetic damage from radon decay progeny may be an important contributor.  This AOP defines a path to cancer using key events  related to DNA damage response and repair. The molecular initiating event (MIE)  which represents the first chemical interaraction with the cell is identified as the direct deposition of ionizing energy.  Energy deposited onto a cell can lead to multiple ionization events to targets such as DNA. This energy will break DNA double strands (KE1) and  initiate  DSB repair machinery.  In higher eukaryotes, this occurs through non-homologous end joining (NHEJ) which is a quick and efficient, but error-prone process (KE2). If DSBs occur in regions of the DNA transcribing critical genes, then mutations (KE3) generated through faulty repair may alter the function of these genes or may cause chromosomal aberrations (KE4), resulting in genomic instability. These events will alter the functions of many gene products and impact cellular pathways such as cell growth, cell cycling, and apoptosis. With these alterations, cell proliferation (KE5) will be promoted by escaping the regulatory control and form hyperplasia in lung epithelial cells, leading eventually to lung cancer (AO) induction and metastasis . The overall weight of evidence for this AOP is strong.   By developing this AOP, we have supported the necessary efforts highlighted by national and international radiation protection agencies to consolidate and enhance the knowledge in understanding the mechanisms of low dose radiation exposures.


Background (optional)

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According to the World Cancer Research Fund, lung cancer is a disease that poses a significant healthcare burden world-wide. (https://www.wcrf.org/dietandcancer/cancer-trends/worldwide-cancer-data).  It is the most commonly diagnosed cancer with the highest incidence of occurrence on a global scale (excluding non-melanoma skin cancers).  It is a multi-faceted disease exhibiting various genetic lesions and involving the accumulation of multiple molecular abnormalities over time.  It is blamed for 1.5 million deaths annually. Although the link between smoking and lung cancer has been well-established, environmental and indoor radiation exposure are also significant contributors.  Risk assessment measures for defining acceptable exposure levels of radiation exposure still remain uncertain; including the scientific research to support the justifications.  This is partially due to the assumption of a non-threshold and linear model at low doses with no consideration that cellular/tissue effects of low dose radiation exposure remain poorly understood.

This AOP has brought together molecular and cellular based research in the radiation realm and  defined a modular, simplistic path towards lung cancer. It has used data–rich key events to a classic targeted response onto a cell that is applicable to multiple radiation stressors (eg. X-rays, gamma rays, alpha particles, beta particles, heavy ions, neutrons) and well supported thorough empirical evidence. Decades of research suggest that energy in the form of ionizing radiation can break DNA molecules.  In vitro mutagenicity studies suggest that alterations in genes in the form of mutations, chromosomal aberrations and micronuclei formation may be important for cancer cell differentiation/proliferation and eventually neoplastic transformation. 

This AOP is also a case example of how existing evidence from radiation stressors can fortify empirical evidence surrounding key events that may be non-radiation specific and vice versa. By using a radiation centric molecular initiating event (MIE), networks can be developed for multiple adverse outcomes distinct to a radiation response.  As different radiation stressors can trigger the MIE, the AOP will have wide applicability. It is our  goal, with the development of this AOP to motivate radiation researchers to use this framework for bringing together research data, exchanging knowledge, identifying  priority areas and better co-ordinating research in the low-dose ionizing radiation field. 


Summary of the AOP

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Events: Molecular Initiating Events (MIE)

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Key Events (KE)

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Adverse Outcomes (AO)

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Sequence Type Event ID Title Short name
1 MIE 1686 Direct Deposition of Energy Energy Deposition
2 KE 1635 Increase, DNA strand breaks Increase, DNA strand breaks
3 KE 155 N/A, Inadequate DNA repair N/A, Inadequate DNA repair
4 KE 185 Increase, Mutations Increase, Mutations
5 KE 1636 Increase, Chromosomal aberrations Increase, Chromosomal aberrations
6 KE 870 Increase, Cell Proliferation Increase, Cell Proliferation
AO 1556 Increase, lung cancer Increase, lung cancer

Relationships Between Two Key Events
(Including MIEs and AOs)

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Title Adjacency Evidence Quantitative Understanding
Energy Deposition leads to Increase, DNA strand breaks adjacent High High
Increase, DNA strand breaks leads to N/A, Inadequate DNA repair adjacent Moderate Moderate
N/A, Inadequate DNA repair leads to Increase, Mutations adjacent Moderate Moderate
N/A, Inadequate DNA repair leads to Increase, Chromosomal aberrations adjacent High Low
Increase, Mutations leads to Increase, Cell Proliferation adjacent High Low
Increase, Chromosomal aberrations leads to Increase, Cell Proliferation adjacent Moderate Low
Increase, Cell Proliferation leads to Increase, lung cancer adjacent High Low
Energy Deposition leads to Increase, Mutations non-adjacent High High
Energy Deposition leads to Increase, Chromosomal aberrations non-adjacent High High
Energy Deposition leads to Increase, lung cancer non-adjacent Moderate Moderate
Increase, DNA strand breaks leads to Increase, Mutations non-adjacent High Low
Increase, DNA strand breaks leads to Increase, Chromosomal aberrations non-adjacent High Low
Increase, Mutations leads to Increase, lung cancer non-adjacent High Low
Increase, Chromosomal aberrations leads to Increase, lung cancer non-adjacent Moderate Moderate

Network View

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Stressors

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Name Evidence Term
Ionizing Radiation High

Life Stage Applicability

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Life stage Evidence
All life stages High

Taxonomic Applicability

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Term Scientific Term Evidence Link
human Homo sapiens High NCBI
rat Rattus norvegicus High NCBI
mouse Mus musculus High NCBI

Sex Applicability

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Sex Evidence
Unspecific High

Overall Assessment of the AOP

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Considerable mechanistic dose-response data has been generated in the radiation field, particularly in the area of clastogenic lesions.  This data has been compiled and captured in this AOP in the most simplified, modular path to lung cancer from a molecular initiating event of deposited energy onto DNA.  This AOP is supported through KERs for which there is biological plausibility and available empirical evidence.  Although it is clear that our proposed AOP is not the only route to the AO, it does represent a classic targeted response of radiation insult on a cell.  The empirical evidence to support this pathway is strong and probabilistic.  As per AOP conventions, the pathway does not describe every mechanism and alteration that is ultimately involved in radiation-associated carcinogenesis. Instead, KEs that are routinely measured using modern and conventional assays are described. For this reason, not all of the KEs that are hallmarks of cancer i.e. evasion, angiogenesis etc. are mapped out, but as they are critical events they can be developed separately. This AOP will be the first to use a MIE that is radiation-specific and therefore can act as a foundational AOP to build networks of radiation-specific responses.  Networks can evolve to multiple AOs with additional KEs that incorporate non-targeted effects, immune and adaptive responses, in parallel.   

 

While this AOP is applicable to other types of radiation-induced cancers, lung cancer was selected as the AO due to its relevance to radon risk assessment and its broader applicability to the chemical field. Lung cancer is a major public health problem world-wide, killing an estimated 1.5 million people annually (https://www.wcrf.org/dietandcancer/cancer-trends/worldwide-cancer-data).  Although smoking is the leading cause of lung cancer, numerous environmental sources are also important contributors including radon, asbestos, air pollution and arsenic (Hubaux et al., 2012).  Some of these stressors can act synergistically to increase risk, particularly among smokers. It has been shown that the histological lung profile of individuals that are smokers is quite different from non-smokers exposed to high radon levels (Egawa et al., 2012).  This is in part due to the complexity of each stressor, in terms of its interaction with cells at the molecular level. As radon is the second leading cause of cancer, distinguishing its mode of action at the cellular level from smoking becomes important. Environmental and indoor radon exposures are significant contributors to lung cancer and risk assessment measures for defining acceptable exposure levels of radon exposure still remain uncertain, including the scientific research to support the justification of these levels (Samet et al., 2000 and 2006).  This is partially due to the assumption of a non-threshold and linear model with no consideration that cellular/tissue effects of low dose radiation exposure remain poorly understood (Ruhm et al., 2016; Shore et al., 2018).

 

Despite the decades of research in the area of radiation and DNA damage, a major challenge in developing this AOP was in finding the required components (i.e. essentiality, temporal, incidence and dose concordance) to provide strong empirical evidence to help support the KERs.  Across all KERs, studies were lacking that used of a broad dose-range.  Most studies conducted analysis at one time-point and there were limited studies that supported the essentiality criteria.  This was particularly evident for the KERs of inadequate repair to mutations/CA and mutations/CA to cellular proliferation. The non-adjacent KERs (i.e. DDOE to CA or DDOE to mutations), were generally more well supported.  Furthermore, no single study encompassed all the KERs proposed in this AOP.  In addition, there were considerable discordant results across KE simply due to the MIE as its outcome is dependent on factors such as cell type, dose, dose-rate, and radiation quality. These factors can influence the amount and type of damage, which in turn can affect the probability to drive a path forward to cancer. The principle knowledge gap arose from the lack of data in the form of essentiality studies, using inhibitors and knock-in genes as well for a number of KERs, there was minimal dose-response and temporal response data in well-conducted animal studies. There is also a range of uncertainty on how confounders such as lifestyle, health status, and radiosensitivities affect an individual’s path to an AO.  Additional KEs may need to be added in parallel as our knowledge in these areas becomes better understood. These challenges can drive research priorities in the future.

An overall assessment of this AOP shows that there is strong biological plausibility and moderate empirical evidence to suggest a qualitative link between deposition of energy on DNA to the final AO of lung cancer. This evidence has been derived predominately from decades of research using laboratory studies and through mathematical simulations of cell-based models.  These studies have shown both dose- and temporal-response relationships for select KEs. The quantitative thresholds to initiate each of the KEs have been shown to vary with factors such as the cell type, dose-rate of exposure and radiation quality. Thus, an absolute amount of deposited energy (MIE) to drive a KE forward to a path of cancer is not yet definable. This is particularly relevant to low doses and low dose-rates of radiation exposure where the biology is interplayed with conflicting concepts of hormesis, hypersensitivity and the linear no threshold theory. Furthermore, due to the stochastic nature of the MIE, it remains difficult to identify specific threshold values of DSBs needed to overwhelm the DNA repair machinery to cause “inadequate” DNA repair leading to downstream genetic abnormalities and eventually cancer. With a radiation stressor, a single hit to the DNA molecule could drive a path forward to lung cancer; however this is with low probability. Conversely, at much higher doses, a cell will induce apoptosis and may not be driven to cancer induction.  Although empirical modeling of cancer probability vs. mean radiation dose and time to lethality, does provide a good visualization of the effective thresholds (Raabe et al., 2011), practically, there is still considerable uncertainty surrounding the connection of biologically contingent observations and stochastic energy deposition. Future work may focus on developing more precise quantitative and predictive models to help address these types of uncertainties. 

This foundational AOP will initiate the building of networks, feedback loops that will further the essential events towards lung cancer, including genome alterations, oxidative stress, and metabolomics effectors.  This will require efforts from the larger radiation community.  As the empirical evidence to support these areas becomes stronger, a better representation of events to lung cancer will emerge. By identifying uncertainties and inconsistencies in the literature, research can be directed to address knowledge gaps, which can later help refine the pathway.  It is our goal, with this AOP to motivate radiation researchers to use this framework for bringing together research data, exchanging knowledge and identifying research priority areas in the low-dose ionizing radiation field.  Long-term, this AOP alongside others in the radiation field will help to identify key events common to chemical stressors and multiple adverse outcomes, which will be important to help refine risk assessment. In all, by building more radiation-relevant AOPs, the AOP framework will have a bigger role in supporting radiation practice.

Domain of Applicability

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This AOP is relevant to mammals (Eymin & Gazzeri, 2009; Barron et al., 2014; Kurgan et al., 2017). The pathway leading to the development of lung cancer often occurs during adulthood but may be applicable at earlier life stages (Liu et al., 2015) and is independent of sex. In humans, however, genetic abnormalities/mutations suggestive of lung cancer risk seem to be influenced by ethnicity (Lloyd et al., 2013), smoking history (Lim et al., 2009; Sanders & Albitar, 2010; Paik et al., 2012; Lloyd et al., 2013; Cortot et al., 2014; Minina et al., 2017), age (Lloyd et al., 2013), sex (Lim et al., 2009; Cortot et al., 2014) and genotype (Lim et al., 2009; Sanders & Albitar, 2010; Kim et al., 2012; Paik et al., 2012; Cortot et al., 2014; Minina et al., 2017). Evidence supporting this AOP comes primarily from studies using bacterial DNA (Sutherland et al., 2000; Jorge et al., 2012), human fibroblast cells (Rothkamm & Lo, 2003; Kuhne et al., 2005; Rydberg et al., 2005a), mice (Duan et al., 2008; Zhang & Jasin, 2011), hamsters (Bracalente et al., 2013; Lin et al., 2014), lung cancer cell lines (Sato, Melville B. Vaughan, et al. 2006; Kurgan et al., 2017; Tu et al., 2018), and tissue samples (both with and without lung cancer) Sun et al., 2016; Tu et al., 2018 Warth et al., 2014.


Essentiality of the Key Events

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Support for Essentiality of KEs

Defining Question

Strong

Moderate

Weak

Are downstream KEs and/or the AO prevented if an upstream KE is blocked?

Direct evidence from specifically designed experimental studies illustrating essentiality for at least one of the important KEs

Indirect evidence that sufficient modification of an expected modulating factor attenuates or augments a KE

No or contradictory experimental evidence of the essentiality of any of the KEs

MIE:

Direct Deposition of Energy

Evidence for Essentiality of KE: Weak

This event is difficult to test for essentiality as deposition of energy is a physical stressor and cannot be blocked/decreased using chemicals.  However, there are a number of antioxidant studies demonstrating that treatment with various antioxidants prior to irradiation decreases the number of radiation-induced DSBs (results summarized in a review by Kuefner et al. 2015; Smith et al. 2017).

KE1:

Double-Strand Breaks, Increase

Evidence for Essentiality of KE: Weak

A variety of different studies demonstrate that organisms with compromised DNA repair tend to have an increased incidence of DSBs. Inhibition studies have shown that addition of a DNA repair antagonist results in significant increases in DSBs at 6 and 12 hours post-irradiation (Dong et al. 2017). Similarly, knock-outs/knock-downs of DNA repair proteins also results in persisting DSBs post-irradiation (Rothkamm and Lo 2003; Bracalente et al. 2013; Mcmahon et al. 2016; Dong et al. 2017), with one DNA ligase IV-deficient human cell line showing DSBs 240 - 340 hours after radiation exposure (Mcmahon et al. 2016). Studies by Tatsumi-Miyajima et al., (1993) note the increased rate of supF mutation frequencies following the use of a restriction, Aval, which induces DSBs in different human fibroblast cell lines transfected with plasmids containing the Aval restriction site.  Kurashige et al. (2017) have demonstrated a decrease in MN frequency following the reduction in DSBs by regulating NAC pre-treatment.

KE2: Inadequate DNA Repair, Increase

Evidence for Essentiality of KE: Strong

There is extensive evidence to demonstrate the essentiality of inadequate repair to downstream events. Studies show that inhibiting DNA repair results in a lack of DNA repair foci post-irradiation (Paull et al. 2000), while cells deficient in ATM (involved in DNA repair) show increased levels of incorrectly rejoined DSBs (Lobrich et al. 2000). Similarly, chromosomal aberrations were more frequent after inhibition of various proteins involved in DNA repair (Chernikova et al. 1999; Heterodimer et al. 2002; Wilhelm et al. 2014). Furthermore, when knock-out cell lines (i.e., knock-out of genes involved in DNA repair to increase the incidence of ‘inadequate’ repair)  were examined for genomic abnormalities, increased incidence of chromosomal aberrations were clearly evident (Karanjawala et al. 1999; Cornforth and Bedford 1994; Patel et al. 1998; Simsek and Jasin 2010; Lin et al. 2014; Wilhelm et al. 2014; Mcmahon et al. 2016).  Deficiencies in proteins involved in DNA repair also resulted in altered mutation frequencies relative to wild-type cases (Amundson and Chen 1996; Feldmann et al. 2000; Smith et al. 2003; Wessendorf et al. 2014; Perera et al. 2016). Mutation frequency increased following knocked-down BER-initiating glycosylases (OGG1, NEIL1, MYH, NTH1) in HEK293T human embryonic kidney cells transfected with plasmids that were either positive or negative for 8-oxodG (Suzuki et al., 2010). Moreoever, G:C to T:A transversion frequency increased in all analyzed cells. Nallanthighal et al. (2017) demonstrated that inadequate DNA repair impacts MN induction in irradiated Ogg1-deficienct mice (compared to Off1+/+ mice).

KE3: Mutations, Increase

Evidence for Essentiality of KE: Strong

Numerous studies show a strong correlation between inadequate DNA repair and mutation incidence, as altered mutation frequencies were evident when there were deficiencies in the proteins involved in DNA repair (Amundson and Chen 1996; Feldmann et al. 2000; Smith et al. 2003; Wessendorf et al. 2014; Perera et al. 2016). Mutations in several different genes, including tumour suppressor gene TP53, have also been shown to increase cell proliferation rates (Hundley et al. 1997; Lang et al. 2004; Ventura et al. 2007; Welcker and Clurman 2008; Duan et al. 2008; Geng et al. 2017; Li and Xiong 2017); mutant or absent TP53 has likewise been implicated in carcinogenesis (Iwakuma and Lozano 2007; Muller et al. 2011; Kim and Lozano 2018). In terms of lung cancer specifically, there are many different studies showing that mutations in TP53, KRAS, and EGFR  are associated with lung carcinogenesis. The conceptual ‘removal’ or ‘blocking’ of these mutations using conditional knock out models, inducible mutation models, and treatment with various antagonizing and agonizing compounds has been observed to reverse or prevent lung tumourigenesis in vivo (Roth et al. 1996; Fisher et al. 2001; Ventura et al. 2007; Iwakuma and Lozano 2007; Jia et al. 2016; Luo et al. 2019, Krasinski 2012). The lung tumourigenesis process was also observed to be expedited by exposure of Gprc5a knock-out mice to a known pulmonary carcinogen; this resulted in more somatic mutations and an increased tumour burden in a much shorter time frame relative to unexposed mice (Fujimoto et al. 2017).   

KE4: Chromosomal Aberrations, Increase

Evidence for Essentiality of KE: Weak

Many studies using a model with inadequate DNA repair (in the form of knock-out cell lines and DNA repair inhibitor studies) demonstrated that chromosomal aberrations were significantly increased when DNA repair was inadequate (Karanjawala et al.; Patel et al. 1998; Deniz Simsek and Jasin 2010; Lin et al. 2014; Wilhelm et al. 2014; Mcmahon et al. 2016, Cornforth 1994). The presence of chromosomal aberrations, particularly gene fusions and translocations, has also been associated with high rates of cellular proliferation (Li et al. 2007; Soda et al. 2007; Guarnerio et al. 2016).There also is support for the essentiality of CAs in the induction of cancer. There were significant increases in CAs (micronuclei, nucleoplasmic bridges and nuclear buds) in peripheral blood lymphocyte cultures after addition of a known pulmonary carcinogen to the cells (Lloyd et al. 2013). Furthermore, introduction of the BCR/ABL translocation in mice resulted in chronic myelogenous leukemia; this was accomplished by lethally irradiating the mice and performing a bone marrow transplant with cells that contained a retrovirus carrying the BCR/ABL translocation (Pear et al. 1998). Furthermore, tumour-inducing A549 cells, which are deficient in TSCL1 due to a loss of heterozygosity at chromosome 11, can induce detectable tumours within 3 weeks of injection; transfection of these A549 cells with genes to correct the TSCL1 deficiency and subsequent injection into mice results in fewer and slower-growing tumours (Kuramochi et al. 2001).

KE5:

Cell Proliferation, Increase

Evidence for Essentiality of KE: Strong

Rates of cellular proliferation have been shown to be increased when there are mutations in key genes associated with cell cycle control, including tumour suppressor gene TP53 (Hundley et al. 1997; Lang et al. 2004; Ventura et al. 2007; Welcker and Clurman 2008; Duan et al. 2008; Geng et al. 2017; Li and Xiong 2017). Cells transformed with various oncogenic mutations that suppressed tumour suppressor genes and enhanced activity of proto-oncogenes also showed increased cellular proliferation rates in the form of higher tumour volumes (Sato et al. 2017). Addition of inhibitors that blocked the pro-proliferative signaling pathway associated with KRAS and EGFR in these oncogenically-transformed cells resulted in lower rates of cellular proliferation (Sato et al. 2017). Similarly, several specific chromosomal gene fusions and translocations have also been associated with increasing the rate of cellular proliferation (Li et al. 2007; Soda et al. 2007; Guarnerio et al. 2016). In cancer cells known to harbor the Philadelphia chromosome (a translocation heavily implicated in the pathogenesis of acute lymphoblastic leukemia), addition of an ERB inhibitor resulted in decreased cellular proliferation rates in the cancer cells (Irwin et al. 2013). In another experiment where human ovarian cancer cells were treated with estrogen, there was an increase in the levels of micronuclei and a corresponding increase in the proliferation rates; addition of an antagonist maintained micronuclei frequencies and cell proliferation rates at control cell levels (Stopper et al. 2003). Cellular proliferation rates were decreased using both in vitro and in vivo carcinogenic models exposed to anti-cancer compounds, which highlights the importance of high cellular proliferation for carcinogenesis (Kassie et al. 2008; Lv et al. 2012; Wanitchakool et al. 2012; Pal et al. 2013; Warin et al. 2014; Tu et al. 2018). Genetic manipulations of genes involved in proliferation also resulted in modified cellular proliferation rates (Lv et al. 2012; Sun et al. 2016).

 


Evidence Assessment

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Support for Biological Plausibility of KERs

Defining Question

Strong

Moderate

Weak

Is there a mechanistic relationship between KEup and KEdown consistent with established biological knowledge?

Extensive understanding of the KER based on extensive previous documentation and broad acceptance; Established mechanistic basis

KER is plausible based on analogy to accepted biological relationships, but scientific understanding is  not completely established

There is empirical support for  statistical association between KEs, but the structural or functional relationship between them is not understood

Direct Deposition of Energy (MIE)    --> Double-Strand Breaks, Increase (KE1)

Evidence for Biological Plausibility of KER: Strong

It is well established that ionizing radiation can cause various types of DNA damage including single-strand and double-strand breaks (DSBs) (reviewed in Lomax et al. 2013). In particular, there is evidence for the direct deposition of energy and a resulting increase in DSBs (Ward 1988; Terato and Ide 2005; Goodhead 2006; Hada and Georgakilas 2008; Okayasu 2012; Lomax et al. 2013; Moore et al. 2014; Desouky et al. 2015; Sage and Shikazono 2017,Asaithamby and Chen, 2011). Structural damage from the deposited energy can induce chemical modifications in the form of breaks to the phosphodiester backbone of both strands of the DNA. (Joiner 2009). DSBs are also often formed by indirect interactions with radiation through water molecules. Energy deposited on water molecules by radiation results in the production of reactive oxygen species that can then damage the DNA (Ward 1988; Desouky et al. 2015; Maier et al. 2016).

Direct Deposition of Energy (MIE)    --> Mutations, Increase (KE3)

Evidence for Biological Plausibility of KER: Strong

Many studies across a variety of different models provide evidence that direct deposition of energy by ionizing radiation results in increased mutation frequencies (Russell et al. 1957; Winegar et al. 1994; Gossen et al. 1995; Suzuki and Hei 1996; Albertini et al. 1997; Dubrova et al. 1998; Dubrova et al. 2000; Canova et al. 2002; Dubrova et al. 2002; Dubrova and Plumb 2002; Masumura et al. 2002; Somers et al. 2004; Burr et al. 2007; Ali et al. 2012; Adewoye et al. 2015; Wilson et al. 2015; Bolsunovsky et al. 2016; Mcmahon et al. 2016; Matuo et al. 2018; Nagashima et al. 2018). Radiation-specific mutational signatures have been identified in a variety of radiation-induced tumours (Sherborne et al. 2015; Behjati et al. 2016), and there is extensive evidence that radiation increases germline mutations in both mice (Dubrova et al. 1998; Dubrova et al. 2000; Dubrova et al. 2002; Somers et al. 2004; Barber et al. 2009; Ali et al. 2012; Adewoye et al. 2015; Wilson et al. 2015) and humans (Dubrova et al. 2002; Dubrova and Plumb 2002).

Direct Deposition of Energy (MIE)    --> Chromosomal Aberrations, Increase (KE4)

Evidence for Biological Plausibility of KER: Strong

Extensive and diverse data from human, animal and in vitro-based studies show ionizing radiation induces a rich variety of chromosomal aberrations (Schmid et al. 2002; Thomas et al. 2003; Maffei et al. 2004; Tucker et al. 2005a; Tucker et al. 2005b; George et al. 2009; Meenakshi and Mohankumar 2013; Santovito et al. 2013; Arlt et al. 2014; Balajee et al. 2014; Han et al. 2014; Vellingiri et al. 2014; Suto et al. 2015; Adewoye et al. 2015; Cheki et al. 2016; Mcmahon et al. 2016; Morishita et al. 2016; Qian et al. 2016; Basheerudeen et al. 2017; Meenakshi et al. 2017; Abe et al. 2018; Jang et al. 2019).The mechanism leading from direct deposition of energy to chromosomal aberrations has been described in several reviews (Smith et al. 2003; Christensen 2014; Sage and Shikazono 2017). Other evidence derives from studies examining the mechanism of copy number variant formation (Arlt et al. 2014) and induction of radiation-induced chromothripsis (Morishita et al. 2016).

Double-Strand Breaks, Increase (KE1) --> Inadequate DNA Repair, Increase (KE2)

Evidence for Biological Plausibility of KER: Strong

 It is well recognized that almost all types of DNA lesions will result in recruitment of repair enzymes and factors to the site of damage, and the pathway involved in the repair of DSBs has been well-documented in a number of reviews, many of which also discuss the error-prone nature of DNA repair (Van Gent et al. 2001; Hoeijmakers 2001a; Khanna and Jackson 2001; Lieber et al. 2003; San Filippo et al. 2008; Lieber et al. 2010; Polo and Jackson 2011; Schipler and Iliakis 2013; Vignard et al. 2013; Betermier et al. 2014; Mehta and Haber 2014; Moore et al. 2014; Rothkamm et al. 2015; Jeggo and Markus 2015; Chang et al. 2017; Sage and Shikazono 2017) Error-prone repair processes are particularly important when DSBs are biologically induced and repaired during V(D)J recombination of developing lymphocytes(Jeggo et al. 1995; Malu et al. 2012) and during meiotic divisions to generate gametes (Murakami and Keeney 2008).

Inadequate DNA Repair, Increase (KE2) --> Mutations, Increase (KE3)

Evidence for Biological Plausibility of KER: Strong

Decades of research have shown that DNA repair pathways are error prone and can cause mutations inherently (such as the error-prone NHEJ) (Sishc and Davis 2017). This error-prone repair, however, may be due more to the structure of the DSB ends rather than the repair machinery; more complex breaks require more processing, increasing the likelihood that there will be errors in the DNA sequence upon completion of repair (Betermier et al. 2014; Waters et al. 2014). After being exposed to ionizing radiation, approximately 25 – 50% of double-strand breaks have been shown to be incorrectly repaired (Löbrich et al. 1998; Kuhne et al. 2000; Lobrich et al. 2000).

Inadequate DNA Repair, Increase (KE2) --> Chromosomal Aberrations, Increase (KE4)

Evidence for Biological Plausibility of KER: Strong

DSBs are repaired by NHEJ and HR. HR uses a template DNA strand to repair DNA damage, while the more error-prone NHEJ simply religates broken ends back together without the use of a template (van Gent et al. 2001; Hoeijmakers 2001; Jeggo and Markus 2015; Sishc and Davis 2017). Chromosomal aberrations may result if DNA repair is inadequate, meaning that the double-strand breaks are misrepaired or not repaired at all (Bignold, 2009; Danford, 2012; Schipler & Iliakis, 2013). A multitude of different types of chromosomal aberrations can occur, depending on the timing and type of erroneous repair. Examples of chromosomal aberrations include copy number variants, deletions, translocations, inversions, dicentric chromosomes, nucleoplasmic bridges, nuclear buds, micronuclei, centric rings, and acentric fragments. A multitude of publications are available that provide details on how these various chromosomal aberrations are formed in the context of inadequate repair (Ferguson and Alt 2001; Venkitaraman 2002; Povirk 2006; Weinstock et al. 2006; Denis Simsek and Jasin 2010; Lieber et al. 2010; Fenech and Natarajan 2011; Danford 2012; Schipler and Iliakis 2013; Mizukami et al. 2014; Russo et al. 2015; Leibowitz et al. 2015; Rode et al. 2016; Vodicka et al. 2018).  

Mutations, Increase (KE3) -->  Cell Proliferation, Increase (KE5)

Evidence for Biological Plausibility of KER: Strong

It is clearly documented that when enough mutations accumulate in critical genes associated with cell cycling or proliferation, there is potential for uncontrollable cell proliferation to occur, which in some cases leads to carcinogenesis (Bertram 2001; Vogelstein and Kinzler 2004; Panov 2005, Lee and Muller 2010). In fact, one of the hallmarks of cancer is sustained proliferative signalling, and one of the enabling characteristics of this increased proliferation is genomic instability/mutations (Hanahan and Weinberg 2011). Thus mutations are particularly dangerous if they occur in proteins controlling the cell cycle checkpoint for entry into proliferation, such as RB and p53 (, Lee and Muller 2010). Activating mutations in proto-oncogenes (Bertram 2001; Vogelstein and Kinzler 2004; Larsen and Minna 2011) Lee and Muller 2010, inactivating mutations in tumour suppressor genes (Bertram 2001; Vogelstein and Kinzler 2004,Lee and Muller 2010) and inactivating mutations in caretaker/stability genes (Vogelstein and Kinzler 2004; Hanahan and Weinberg 2011) are all associated with abnormal increases the rate of cellular proliferation.

Chromosomal Aberrations, Increase (KE4) --> Cell Proliferation, Increase (KE5)

Evidence for Biological Plausibility of KER: Strong

Chromosomal aberrations are formed when there is inadequate DNA repair (Bignold 2009; Danford 2012; Schipler and Iliakis 2013) or errors during mitosis (Levine and Holland 2018). Chromosomal aberrations have been shown to increase cell proliferation when the aberrations result in the activation of proto-oncogenes (Bertram 2001; Vogelstein and Kinzler 2004), the inactivation of tumour suppressor genes (Bertram 2001; Vogelstein and Kinzler 2004),, or the modification of caretaker/stability genes (Vogelstein and Kinzler 2004). Reviews documenting the contribution of CAs to cellular proliferation and/or cancer development (which implies high rates of cellular proliferation) are available (Mes-Masson and Witte 1987; Bertram 2001; Vogelstein and Kinzler 2004; Ghazavi et al. 2015; Kang et al. 2016). The link between chromosomal instability (CIN), which describes the rate of chromosome gains and losses, and cancer development has also been reviewed (Thompson et al. 2017; Gronroos 2018; Targa and Rancati 2018; Lepage et al. 2019).

Cell Proliferation, Increase (KE5) -->  Lung Cancer, Increase (AO)

Evidence for Biological Plausibility of KER: Strong

The means by  which dysregulation of cell proliferation promotes the transformation of normal to carcinogenic cells has been heavily reviewed (Pucci et al. 2000; Bertram 2001; Panov 2005; Eymin and Gazzeri 2009; Hanahan and Weinberg 2011; Larsen and Minna 2011). The cell cycle is essential in controlling cellular proliferation rates, and requires a series of checkpoints to be passed before the cell can fully commit to the process of cell division (Pucci et al. 2000; Bertram 2001; Eymin and Gazzeri 2009; Hanahan and Weinberg 2011). One of the most important checkpoints requires the proper functioning of p53, RB, CDK4 and CDK6. The tumour suppressor p53  plays a particularly important role in stopping the cell cycle when there is DNA damage, and for triggering apoptosis when damage is too severe to be repaired (Bertram 2001; Hanahan and Weinberg 2011; Larsen and Minna 2011). Telomeres also play a role in controlling cell proliferation; when the telomeres become too short to protect the coding DNA, the cell enters into a state of replicative senescence (Bertram 2001; Hanahan and Weinberg 2011). All of these processes play a role in controlling the rate of cellular proliferation within a cell. Cancer may occur when these processes became dysregulated such that cells begin to proliferate at excessively high rates. High rates of proliferation are in fact one of the strongest hallmarks of cancer (Hanahan and Weinberg 2011), and uncontrolled proliferation can be accomplished through sustained proliferative signalling through activation of proto-oncogenes (Bertram 2001; Vogelstein and Kinzler 2004; Eymin and Gazzeri 2009; Hanahan and Weinberg 2011; Larsen and Minna 2011), evading growth suppressors and resisting cell death through suppression of tumour suppressor genes (Bertram 2001; Vogelstein and Kinzler 2004; Eymin and Gazzeri 2009; Hanahan and Weinberg 2011; Larsen and Minna 2011), and overcoming replicative senescence through expression of the telomere-lengthening enzyme telomerase (Bertram 2001; Panov 2005; Hanahan and Weinberg 2011; Larsen and Minna 2011). In lung cancer specifically, commonly activated proto-oncogenes include EGFR, ERBB2, MYC, KRAS, MET, CCND1, CDK4 and BCL2, while commonly inactivated tumour suppressor genes are TP53, RB1, STK11, CDKN2A, FHIT, RASSF1A, and PTEN (Larsen and Minna 2011). Telomerase is also activated in nearly all small cell lung cancer (SCLC) cases, and in over three-quarters of non-small cell lung cancer (NSCLC) cases (Panov 2005; Larsen and Minna 2011).

 

Double-Strand Breaks, Increase (KE1) --> Mutations, Increase (KE3)

Evidence for Biological Plausibility of KER: Strong

Mechanisms of DNA strand break repair have been extensively studied. It is accepted that non-homologous joining of broken ends can introduce deletions, insertions, or base substitution. In mamalian and yeast cells, both HR and NHEJ can lead to alteration in DNA sequence (Hicks & Haber, 2010; Butning & Nussenzweig, 2013; Byrne et al., 2014; Rodgers & McVey, 2016; Dwivedi & Haver, 2018).

 

Double-Strand Breaks, Increase (KE1) --> Chromosomal Aberrations, Increase (KE4)

Evidence for Biological Plausibility of KER: Strong

DNA strand breaks must occur for chromosomal aberrations to occur. Studies have shown DSBs leading to irreversible damage. The links between DSBs and the role DSB repairs has in preventing chromosomal aberrations is widely discussed, with several reviews available: (van Gent et al., 2001; Ferguson & Alt, 2001; Hoeijmakers, 2001; Iliakis et al., 2004; Povirik, 2006; Weinstock et al., 2006; Natarajan & Palitti, 2008; Lieber et al., 2010; Mehta & Haber, 2014; Ceccaldi et al., 2016; Chang et al., 2017; Sishc & Davis, 2017; Brunet & Jasin, 2018).

Mutations, Increase (KE3) --> Lung Cancer, Increase (AO)

Evidence for Biological Plausibility of KER: Moderate

There is strong biological plausibility for the relationship between mutations and lung cancer. Bioinformatics studies have identified unique mutation signature profiles associated with specific types of cancer, including lung adenocarcinoma, lung squamous cell carcinoma and lung small cell carcinoma (Alexandrov et al. 2013; Jia et al. 2014). Moreover, mutations/genome instability have been implicated as one of the ‘enabling characteristics’ underlying the hallmarks of cancer (Hanahan and Weinberg 2011). Mutations are thought to promote tumourigenesis by modifying the expression of tumour suppressor genes, proto-oncogenes, and caretaker/stability genes in such a way that promotes cell proliferation and/or suppresses apoptosis (Vogelstein and Kinzler 2004; Panov 2005; Sanders and Albitar 2010; Hanahan and Weinberg 2011; Larsen and Minna 2011).  Commonly mutated genes in lung cancer include TP53, KRAS and EGFR. Mutations in these genes, along with known lung cancer driver mutations, are thought to promote tumourigenesis by stimulating pro-proliferation signalling pathways such as the PI3K-AKT-mTOR pathway and RAS-REF-MEK pathway (Varella-garcia 2009; Sanders and Albitar 2010; Larsen and Minna 2011McCubrey 2006).

Chromosomal Aberrations, Increase (KE4) --> Lung Cancer, Increase (AO)

Evidence for Biological Plausibility of KER: Moderate

Chromosomal aberrations, encompassing chromosome-type aberrations, chromatid-type aberrations, micronuclei, and nucleoplasmic bridges, have all been found to be predictive of cancer risk in various human cohorts (Bonassi et al. 2000; Smerhovsky et al. 2002; Hagmar et al. 2004; Norppa et al. 2006; Boffetta et al. 2007; Bonassi et al. 2008; Lloyd et al. 2013; El-zein et al. 2014; Vodenkova et al. 2015; El-zein et al. 2017). Specific categories of CAs, including CNVs (Wrage et al. 2009; Shlien and Malkin 2009; Liu et al. 2013; Mukherjee et al. 2016; Zhang et al. 2016; Ohshima et al. 2017) and gene rearrangements (Bartova et al. 2000; Trask 2002; Sanders and Albitar 2010; Sasaki et al. 2010; Mao et al. 2011), have also been associated with cancer development. Chromosomal aberrations promote tumourigenesis through the alteration of pathways controlling cellular growth and apoptosis (Albertson et al. 2003; Sanders and Albitar 2010). The chromosomal aberration burden may be increased by factors such as aberrant centromeres, telomerase deficiencies paired with poor cell surveillance (Albertson et al. 2003), ionizing radiation (Hei et al. 1994; Weaver et al. 1997; Weaver et al. 2000), and the interplay between non-clonal and clonal CAs (Heng, Bremer, et al. 2006; Heng, Stevens, et al. 2006).

Direct Deposition of Energy (MIE)    --> Lung Cancer, Increase (AO)

Evidence for Biological Plausibility of KER: Strong

The direct deposition of energy, particularly by radon gas, has been associated heavily with lung cancer (Axelson 1995; Jostes 1996; Beir 1999; Kendall and Smith 2002a; Al-Zoughool and Krewski 2009; Robertson et al. 2013). Deposition of energy that triggers lung carcinogenesis in particular is thought to enter the body through inhalation (Beir 1999; Kendall and Smith 2002b). The inhaled particles are thought to deposit on lung tissue and decay, producing ionizing radiation (Axelson 1995; Beir 1999; Kendall and Smith 2002b; Al-Zoughool and Krewski 2009) that can direct the cell towards carcinogenesis (Axelson 1995; Beir 1999; Robertson et al. 2013). The process of radiation-induced carcinogenesis often follows three steps: initiation, promotion and progression. Initiation refers to the interaction between the radiation and the cell, and results in irreversible genetic changes. Promotion occurs when non-carcinogenic promoter is added to the initiated cells such that it synergistically increases oncogenesis, often through receptor-mediated epigenetic changes. Progression occurs at the point when the cells convert from benign to malignant, and is associated with rapid growth and further accumulation of genomic aberrations (NRC 1990; Pitot 1993).

 

Support for Empirical Evidence of KERs

Defining Question

Strong

Moderate

Weak

Does empirical evidence support that a change in KEup leads to an appropriate change in KEdown? Does KEup occur at lower doses and earlier time points than KEdown and is the incidence of KEup > than that for KEdown?

 

Inconsistencies?

Multiple studies showing dependent change in both events following exposure to a wide range of specific stressors (Extensive evidence for temporal, dose-response and incidence concordance);  No or few critical data gaps or conflicting data

Demonstrated dependent change in both events following exposure to a small number of specific stressors; Some evidence inconsistent with expected pattern that can be explained by factors such as the experimental design, technical considerations, differences between laboratories, etc.

Limited or no studies reporting dependent change in both events following exposure to a specific stressor (i.e. endpoints never measured in the same study or not at all); And/or significant inconsistencies in empirical support across taxa and species that don’t align with expected pattern for hypothesized AOP

Direct Deposition of Energy (MIE) --> Double-Strand Breaks, Increase (KE1)

Evidence for Empirical Support of KER: Strong

Evidence exists for dose/incidence and temporal concordance between deposition of energy and the resultant formation of DNA double-strand breaks. With increasing ionizing radiation, there is an increase in the frequency of double-strand breaks (Charlton et al. 1989; Rogakou et al. 1999; Sutherland et al. 2000; Lara et al. 2001; Rothkamm and Lo 2003; Kuhne et al. 2005; Sudprasert et al. 2006; Rube et al. 2008; Beels et al. 2009; Grudzenski et al. 2010; Flegal et al. 2015; Shelke and Das 2015; Antonelli et al. 2015). However, dose-rate and radiation quality play a crucial role in determining the degree of DNA damage. Temporally, DSBs have been evident at 3 - 30 minutes post-irradiation (Rogakou et al. 1999; Rothkamm and Lo 2003; Rube et al. 2008; Beels et al. 2009; Kuefner et al. 2009; Grudzenski et al. 2010; Antonelli et al. 2015). A significant proportion of the DSBs are resolved within 5 hours of radiation (Rogakou et al. 1999; Rube et al. 2008; Kuefner et al. 2009; Grudzenski et al. 2010; Shelke and Das 2015), with a return to baseline levels by 24 hours in most cases (Rothkamm and Lo 2003; Rube et al. 2008; Grudzenski et al. 2010; Antonelli et al. 2015).

Direct Deposition of Energy (MIE) --> Mutations, Increase (KE3)

Evidence for Empirical Support of KER: Strong

Evidence exists for dose/incidence concordance between deposition of energy by ionizing radiation and a corresponding dose-dependent increase in mutation frequency (Suzuki and Hei 1996; Canova et al. 2002; Bolsunovsky et al. 2016; Mcmahon et al. 2016; Matuo et al. 2018; Nagashima et al. 2018). The linear energy transfer of the radiation (Dubrova and Plumb 2002; Matuo et al. 2018), whether the radiation is chronic or acute (Russell 1958), the radiation type (Masumura 2002), and the tissue being irradiated (Masumura 2002, Gossen 1995) all affect this dose-dependent increase. Temporally, it is well established that an increased incidence of mutations is reported after the deposition of energy by radiation (Winegar 1994, Gossen 1995, Albertini 1997, Dubrova 2002A, Matuo 2018, Canova 2002, Nagashima 2018, Masumura 2002, Russell 1958). Most of these studies, however, span over days and weeks, thus making it difficult to pinpoint exactly when mutations occur. Several studies report the manifestation of mutations within 2 - 3 days of irradiation (Winegar 1994, Masumura 2002, Gossen 1995), with an increased mutation frequency still elevated at 14 (Winegar 1994) and 21 days (Gossen 1995) after radiation exposure.

Direct Deposition of Energy (MIE) -->  Chromosomal Aberrations, Increase (KE4)

Evidence for Empirical Support of KER: Strong

Results from many studies indicate dose/incidence and temporal concordance between the deposition of energy and the increased frequency of chromosomal aberrations. There is strong evidence of a dose-dependent increase in a wide range of chromosomal aberrations in response to increasing radiation dose (Schmid 2002, Thomas 2003, Jang 2019, Abe 2018, Suto 2015, McMahon 2016, Tucker 2005A, Tucker 2005B, Arlt 2014, McMahon 2016, Balajee 2014,George 2009, Maffei 2004, Qian 2015). Temporally, it is well-established that chromosomal aberrations occur after exposure to radiation (Schmid 2002, Thomas 2003, Balajee 2014, Arlt 2014, George 2009, Suto 2015, Basheerudeen 2017, Tucker 2005A, Tucker 2005B, Abe 2018, Jang 2019), though the exact timing is difficult to pinpoint because most assays take place hours or days after the radiation exposure. One notable study did, however, document the presence of chromosomal aberrations within the first 20 minutes of irradiation, with the frequency increasing sharply until approximately 40 minutes, followed by a plateau (McMahon 2016). By 7 days post-irradiation, the frequencies of most chromosomal aberrations had declined (Tucker 2005A, Tucker 2005B).  It should be noted that chromosomal aberrations induced by ionizing radiation are dependent on dose, dose-rate, and radiation type (Bender et al., 1988; Guerrero-Carbajal et al., 2003; Day et al., 2007, Suzuki 1996).  

Double-Strand Breaks, Increase (KE1) --> Inadequate DNA Repair, Increase (KE2)

Evidence for Empirical Support of KER: Moderate

Results from many studies indicate dose/incidence and temporal concordance between the frequency of double-strand breaks and the rate of inadequate repair. As DNA damage accumulates in organisms, the incidence of in adequate DNA repair activity (in the form of non-repaired or misrepaired DSBs) also increases (Dikomey 2000, McMahon 2016, Kuhne 2005, Rydberg 2005, Kuhne 2000, Lobrich 2000). DNA damage and its ensuing repair also follow a very similar time course, with both events documented within minutes of a radiation stressor (Pinto 2005, Rothkamm 2003, Asaithambly 2009, Dong 2017, Paull 2000). Uncertainties in this KER include controversy surrounding how error-prone NHEJ truly is (Betemier 2014), differences in responses depending on the level of exposure of a genotoxic substance (Marples 2004), and confounding factors (such as smoking) that affect double-strand break repair fidelity (Scott 2006, Leng 2008).

Inadequate DNA Repair, Increase (KE2) --> Mutations, Increase (KE3)

Evidence for Empirical Support of KER: Moderate

There are several studies that indicate a dose/incidence concordance between inadequate DNA repair and an increased frequency of mutations. Inadequate DNA repair (Ptácek et al. 2001; Mcmahon et al. 2016) and mutation frequencies (Mcmahon et al. 2016) have both been found to increase in a dose-dependent fashion with increasing doses of a radiation stressor. Moreover, specific genomic regions with inadequate DNA repair rates also were found to have increased mutation densities in cancer samples (Perera et al. 2016). Increased mutation frequencies have also been demonstrated in cases where more complex DNA repair is required (Smith et al. 2001). According to the results of this study, evidence of repaired DNA was present prior to the detection of mutations in cases of simple repair, whereas these two events occurred together at a later time point when more complex repair was required (Smith et al. 2001).

Inadequate DNA Repair, Increase (KE2) --> Chromosomal Aberrations, Increase (KE4)

Evidence for Empirical Support of KER:  Moderate

There is little empirical evidence available that directly examines the dose and incidence concordance between DNA repair and CAs within the same study. However, comparison of results from studies that measure either radiation-induced DNA repair or radiation-induced chromosomal aberrations demonstrate that the rate of double-strand break misrepair increases in a dose-dependent fashion with radiation doses between 0 - 80 Gy (Mcmahon et al. 2016), as does the incidence of chromosomal aberrations between doses of 0 - 10 Gy (Thomas et al. 2003; Tucker et al. 2005a; Tucker et al. 2005b; George et al. 2009; Arlt et al. 2014; Balajee et al. 2014; Han et al. 2014; Suto et al. 2015; Mcmahon et al. 2016). Similarly, there is not clear evidence of a temporal concordance between these two events. One study examining DNA repair and micronuclei in irradiated cells pre-treated with a DNA repair inhibitor found that both repair and micronuclei were present at 3 hours and 24 hours post-irradiation. This suggests that there may be temporal concordance (Chernikova et al. 1999). More research, however, is required to establish empirical evidence for this KER.

Mutations, Increase (KE3)    --> Cell Proliferation, Increase (KE5)

Evidence for Empirical Support of KER: Moderate

There is little empirical evidence available that assesses the dose and incidence concordance between mutation frequency and cellular proliferation rates. The correlation between these two events is clear in human epidemiology studies examining the incidence between mutations in specific genes, such as TP53 and BRCA1, and the proliferative status of human tumours (M Jarvis et al. 1998; Schabath et al. 2016). Another study introducing oncogenic mutations into mouse lung epithelial cells demonstrated that the addition of multiple oncogenic mutations to the cells resulted in increased tumour volumes over 40 days (suggestive of cell proliferation); in contrast, cells containing only one of these mutations did not show significant changes in tumour volumes (Sato et al. 2017). Unsurprisingly, there is also little empirical evidence available supporting a temporal concordance between these two events. One review explores the timing between these two events by comparing the somatic mutation theory of cancer and the stem cell division theory of cancer. In the somatic mutation theory, it is suggested that mutations accumulate and result in increased rates of cellular proliferation; the stem cell theory, however, states that high proliferation in stem cells allows the accumulation of mutations (López-lázaro 2018). More research is thus required to establish empirical evidence for this KER.

 

Chromosomal Aberrations, Increase (KE4)    --> Cell Proliferation, Increase (KE5)

Evidence for Empirical Support of KER: Moderate

There is little empirical evidence available that assesses the dose and incidence concordance between chromosomal aberration frequency and cellular proliferation rates. There are several reviews available that discuss the structure and function of specific human cancer-associated chromosomal aberrations, including BCR-ABL1, ALK fusions, and ETV6-RUNX1 (Mes-Masson and Witte 1987; Ghazavi et al. 2015; Kang et al. 2016). There was no identified evidence supporting dose and incidence concordance. Details from a study where estrogen-responsive cancer cells were treated with estrogen suggested the possibility of a temporal concordance, as both micronuclei levels and proliferation rates were higher in the estrogen-treated cells at 140 and 216 hours post-treatment (Stopper et al. 2003). Overall, however, more empirical evidence is required to support this KER.

 

Cell Proliferation, Increase (KE5)    --> Lung Cancer, Increase (AO)

Evidence for Empirical Support of KER: Moderate

There is some empirical evidence of a dose and incidence concordance between cell proliferation and lung carcinogenesis. In a few experiments, rodent lungs exposed to various carcinogens showed increased levels of proliferation and developed squamous metaplasia (Zhong et al. 2005) or full-blown tumours (Kassie et al. 2008). Furthermore, nude mice injected with carcinogenic human NSCLC cells also developed tumours within a few weeks of the injection (Pal et al. 2013; Warin et al. 2014; Sun et al. 2016; Tu et al. 2018)(Sun 2016, Pal 2013, Tu 2018, Warin 2014). In terms of temporal concordance between these two events, studies are also limited. Multiple tumour xenograft experiments found that nude mice injected with NSCLC cells develop detectable tumours within two weeks of inoculation, which continued to increase in size over time (Sun 2016, Pal 2013, Tu 2018, Warin 2014). Examination of lung squamous metaplasia after 14 weeks of exposure to high levels of tobacco smoke showed increased cell proliferation markers in comparison to lungs from rats exposed to filtered air (Zhong et al. 2005). Similarly, lung tumours from mice that received carcinogens NNK and BaP orally over 4 weeks were also found to express proliferation markers when examined 27 weeks after the start of the experiment (Kassie et al. 2008).

 

 

Double-Strand Breaks, Increase (KE1) --> Mutations, Increase (KE3)

Evidence for Empirical Support of KER: Moderate

There is some evidence demonstrating dose and temporal concordance between the two KEs, both in-viv and in-vitro. These studies used a variety of sources of ionizing radiation as stressors. The types of radiation testing this relationship include X-rays, gamma-rays, alpha particles and heavy ions. Example studies include: (in vitro) Rydberg et al., 2005; Kuhne et al., 2005, 2000; Dikomey et al., 2000; Lobrich et al., 2000, (in vivo) Ptacek et al., 2001. For a discussion of chemical stressors affecting this relationship, see AOP 296.

Double-Strand Breaks, Increase (KE1) --> Chromosomal Aberrations, Increase (KE4)

Evidence for Empirical Support of KER: Moderate

Temporal concordance is clear in both in vitro and in vivo data. However, due to the differences in the methods used to measure strand breaks and chromosomal aberrations, the dose-response of these events often appear to be discordant. Examples of studies relating the links between DSBs and chromosomal aberrations include an in-vitro study of gamma-radiated lymphoblasted cell lines (Trenz et al., 2003) isolated lymphocytes and whole blood samples (Sudpresert et al., 2006) and PL61 cells (Chernikova et al., 1999). Source of high linear energy transfer have also been probed, see Iliakis et al. (2019).

Mutations, Increase (KE3)    --> Lung Cancer, Increase (AO)

Evidence for Empirical Support of KER: Moderate

Evidence for dose/incidence concordance comes from studies with similar radiological and biological conditions that assessed either the relationship between radiation exposure and mutations, or radiation exposure and cancer. Using various in vitro  models, there was a dose-dependent relationship found for mutation induction and radiation dose (Suzuki and Hei 1996; Weaver et al. 1997; Canova et al. 2002), and for oncogenic transformations and radiation dose (Hei et al. 1994; Miller et al. 1995; Miller et al. 1999). Analyses of lung cancer incidences in radon-exposed rats and uranium miners echo these results (Monchaux et al. 1994; Lubin et al. 1995; Ramkissoon et al. 2018). Likewise, administration of a known pulmonary carcinogen to Gprc5a knock-out mice resulted in an increased rate of tumourigenesis and increased mutation accumulation relative to saline-treated mice (Fujimoto et al. 2017). Increasing the number of mutations in vitro  and  in vivo resulted in cells becoming increasingly more oncogenic (Sato, Melville B Vaughan, et al. 2006; Sasai et al. 2011) and mice sporting a faster rate of lung tumourigenesis (Fisher et al. 2001; Kasinski and Slack 2012), respectively. In terms of temporal concordance, there is some evidence from separate studies indication that mutations precede tumourigenesis (Hei et al. 1994; Lubin et al. 1995; Hei et al. 1997; Miller et al. 1999; Fujimoto et al. 2017) , particulary in Cre-inducible models where Cre expression must be induced for the mutations to be expressed (Fisher et al. 2001; Kasinski and Slack 2012).

Chromosomal Aberrations, Increase (KE4)    --> Lung Cancer, Increase (AO)

Evidence for Empirical Support of KER: Moderate

Evidence for dose/incidence concordance comes from epidemiological studies of radon-exposed uranium miners that found there was an increased CA load with increasing radon exposure (Smerhovsky et al. 2002), and an increased risk of lung cancer with increased cumulative radon exposure (Tirmarchel et al. 1993; Smerhovsky et al. 2002; Vacquier et al. 2008; Walsh et al. 2010). In vivo and in vitro studies have also shown a dose-dependent increase in CAs in lung and non-lung cell lines (Nagasawa et al. 1990; Deshpande et al. 1996; Yamada et al. 2002; Stevens et al. 2014) and lung cells of rodents with increasing radiation dose (A.L. Brooks et al. 1995; Khan et al. 1995; Werner et al. 2017), and a dose-dependent increase in oncogenic transformation in non-lung cells lines (Robertson et al. 1983; Miller et al. 1996)  and in rodent lung tumours with increasing radiation dose (Monchaux et al. 1994; Yamada et al. 2017) Furthermore, there are several published reviews that provide evidence for associations between radon exposure and the appearance of CAs, and radon exposure and the incidence of lung cancer (Jostes 1996; Al-Zoughool and Krewski 2009; Robertson et al. 2013). Likewise, more CAs were found to accumulate in larger tumours (To et al. 2011) and in increasingly more oncogenic lung tissue lesions (Thibervile et al. 1995; Wistuba et al. 1999). There is also evidence for temporal concordance as, the time gap between radiation exposure and the increased incidence of CAs is hours to days (Nagasawa et al. 1990; A.A.L. Brooks et al. 1995; Deshpande et al. 1996; Yamada et al. 2002; Stevens et al. 2014; Werner et al. 2017), while the time gap between radiation exposure and the development of oncogenic transformations or lung tumours is weeks, months or years (Robertson et al. 1983; Tirmarchel et al. 1993; Miller et al. 1996; Pear et al. 1998; Kuramochi et al. 2001; Yamada et al. 2017).

Direct Deposition of Energy (MIE)  --> Lung Cancer, Increase (AO)

Evidence for Empirical Support of KER: Moderate

There is strong evidence of the relationship between radiation exposure and lung carcinogenesis in human epidemiological studies that assess radon exposure and the risk of lung cancer. Results from numerous studies assessing indoor residential radon exposure and outdoor radon exposure in miners suggest that there is a positive association between cumulative radon exposure and lung cancer risk (Darby et al. 2005; Krewski et al. 2005) (Krewski et al. 2006; Torres-Durán et al. 2014; Sheen et al. 2016; Lubin et al. 1995; Hazelton et al. 2001; Al-Zoughool and Krewski 2009; Rodríguez-Martínez et al. 2018; Ramkissoon et al. 2018). Several in vitro studies showed that cells could be induced to obtain oncogenic characteristics through radiation exposure (Hei et al. 1994; Miller et al. 1995). Likewise, irradiation of rats at radon levels comparable to those experienced by uranium miners resulted in a dose-dependent increase in lung carcinoma incidence (Monchaux et al. 1994). There is also evidence of temporal concordance, as the oncogenic characteristics of the radon-exposed cells were not evident until weeks after the irradiation (Hei et al. 1994; Miller et al. 1995), while tumours took months to years to grow (Hei et al. 1994; Monchaux et al. 1994). In humans, the risk of lung cancer was also found to increase with increasing time since exposure (Hazelton et al. 2001) and with longer periods of exposure (Lubin et al. 1995).

 


Quantitative Understanding

?

There is strong biological plausibility and empirical evidence to suggest a qualitative link between deposition of energy on DNA to the final adverse outcome of lung cancer. This evidence has been derived predominately from laboratory studies and through mathematical simulations using cell-based models. The studies show both dose and temporal-response relationships for a select KEs. The quantitative thresholds to initiate each of the KEs are not definitive and have been shown to vary with factors such as the cell type, dose-rate of exposure and radiation quality. Thus, an absolute amount of deposited energy (MIE) to drive a key event forward to a path of cancer is not yet definable. This is particularly relevant to low doses and low dose-rates of radiation exposure where the biology is interplayed with conflicting concepts of hormesis, hypersensitivity and the linear no threshold theory. Furthermore due to the stochastic nature of the MIE, it remains difficult to identify specific threshold values of DSBs needed to overwhelm the DNA repair machinery to cause “inadequate” DNA repair leading to downstream genetic abnormalities and eventually cancer. With a radiation stressor, a single hit to the DNA molecule could drive a path forward to lung cancer; however this is with low probability.  Empirical modeling of cancer probability vs. mean radiation dose and time to lethality, does provide a good visualization of the effective thresholds (Raabe 2011). However, in general there is considerable uncertainty surrounding the connection of biologically contingent observations and stochastic energy deposition.

Raabe OG. Toward improved ionizing radiation safety standards. Health Phys 101: 84–93; 2011.

Support for Quantitative Understanding of KERs

Defining Question

Strong

Moderate

Weak

What is the extent to which a change in KEdown can be predicted from KEup? What is the precision with which uncertainty in the prediction of KEdown can be quantified? What is the extent to which known modulating factors or feedback mechanisms can be accounted for? What is the extent to which the relationships can be reliably generalized across the applicability domain of the KER?

Change in KEdown can be precisely predicted based on a relevant measure of KEup; Uncertainty in the quantitative prediction can be precisely estimated from the variability in the relevant KEup measure; Known modulating factors and feedback/ feedforward mechanisms are accounted for in the quantitative description; Evidence that the quantitative relationship between the KEs generalizes across the relevant applicability domain of the KER

Change in KEdown can be precisely predicted based on relevant measure of KEup; Uncertainty in the quantitative prediction is influenced by factors other than the variability in the relevant KEup measure; Quantitative description does not account for all known modulating factors and/or known feedback/ feedforward mechanisms; Quantitative relationship has only been demonstrated for a subset of the overall applicability domain of the KER

Only a qualitative or semi-quantitative prediction of the change in KEdown can be determined from a measure of KEup; Known modulating factors and feedback/ feedforward mechanisms are not accounted for; Quantitative relationship has only been demonstrated for a narrow subset of the overall applicability domain of the KER

Direct Deposition of Energy (MIE) --> Double-Strand Breaks, Increase (KE1)

Evidence for Quantitative Understanding of KER: Strong

The vast majority of studies examining energy deposition and incidence of DSBs suggest a positive, linear relationship between these two events (Sutherland et al. 2000; Lara et al. 2001; Rothkamm and Lo 2003; Kuhne et al. 2005; Rube et al. 2008; Grudzenski et al. 2010; Shelke and Das 2015; Antonelli et al. 2015). Predicting the exact number of DSBs from the deposition of energy, however, appears to be highly dependent on the biological model, the type of radiation and the radiation dose range, as evidenced by the differing calculated DSB rates across studies (Charlton et al. 1989; Rogakou et al. 1999; Sutherland et al. 2000; Lara et al. 2001; Rothkamm and Lo 2003; Kuhne et al. 2005; Rube et al. 2008; Grudzenski et al. 2010; Antonelli et al. 2015) .

Direct Deposition of Energy (MIE) --> Mutations, Increase (KE3)

Evidence for Quantitative Understanding of KER: Strong

Most studies indicate a positive, linear relationship between the radiation dose and the mutation frequency (Russell et al. 1957; Albertini et al. 1997; Canova et al. 2002; Dubrova et al. 2002; Nagashima et al. 2018). In order to predict the number of mutations induced by a particular dose of radiation, parameters such as the type of radiation, the radiation’s LET, and the type of model system being used should be taken into account (Albertini et al. 1997; Dubrova et al. 2002; Matuo et al. 2018; Nagashima et al. 2018). Predicting the mutation frequency at particular time-points, however, would be very difficult owing to our limited time scale knowledge.

Direct Deposition of Energy (MIE) -->  Chromosomal Aberrations, Increase (KE4)

Evidence for Quantitative Understanding of KER: Strong

Most studies indicate a positive, linear-quadratic relationship between the deposition of energy by ionizing radiation and the frequency of chromosomal aberrations (Schmid et al. 2002; Suto et al. 2015; Abe et al. 2018; Jang et al. 2019). Equations describing this relationship were given in a number of studies (Schmid et al. 2002; George et al. 2009; Suto et al. 2015; Abe et al. 2018; Jang et al. 2019), with validation of the dose-response curve performed in one particular case (Suto et al. 2015). In terms of time scale predictions, this may still be difficult owing to the often-lengthy cell cultures required to assess chromosomal aberrations post-irradiation. For translocations in particular, however, one study defined a linear relationship between time and translocation frequency at lower radiation doses (0 - 0.5 Gy) and a linear quadratic relationship at higher doses (0.5 - 4 Gy) (Tucker et al. 2005b).

Double-Strand Breaks, Increase (KE1)  --> Inadequate DNA Repair, Increase (KE2)

Evidence for Quantitative Understanding of KER: Moderate

According to studies examining DSBs and DNA repair after exposure to a radiation stressor, there was a positive linear relationship between DSBs and radiation dose (Lobrich et al. 2000; Rothkamm and Lo 2003; Kuhne et al. 2005; Asaithamby and Chen 2009), and a linear-quadratic relationship between the number of misrejoined DSBs and radiation dose (Kuhne et al. 2005) which varied according to LET (Rydberg et al. 2005b) and dose-rate (Dikomey and Brammer 2000) of the radiation. Overall, 1 Gy of radiation may induce between 35 and 70 DSBs (Dubrova et al. 2002; Rothkamm and Lo 2003), with 10 - 15% being misrepaired at 10 Gy (Mcmahon et al. 2016) and 50 - 60% being misrepaired at 80 Gy (Lobrich et al. 2000; Mcmahon et al. 2016). Twenty-four hours after radiation exposure the frequency of misrepair appeared to remain relatively constant around 80%, a rate that was maintained for the next ten days of monitoring (Kuhne et al. 2000).

Inadequate DNA Repair, Increase (KE2) --> Mutations, Increase (KE3)

Evidence for Quantitative Understanding of KER: Moderate

Positive relationships have been reported between radiation stressor and inadequate DNA repair, radiation stressor and mutation frequency (Mcmahon et al. 2016), and inadequate DNA repair and mutation frequency (Perera et al. 2016). It has been found that 10 - 15% of DSBs are misrepaired at 10 Gy (Mcmahon et al. 2016) and 50 - 60% at 80 Gy (Lobrich et al. 2000; Mcmahon et al. 2016), with mutation rates varying from 0.1 - 0.2 mutation per 104 cells at 1 Gy and 0.4 - 1.5 mutation per 104 cells at 6 Gy (Mcmahon et al. 2016).

Inadequate DNA Repair, Increase (KE2) --> Chromosomal Aberrations, Increase (KE4)

Evidence for Quantitative Understanding of KER: Weak

A direct quantitative understanding of the relationship between inadequate DNA repair and chromosomal aberrations has not been established. However, some data has been generated using studies from radiation stressor studies. At a radiation dose of 10 Gy, the rate of DSB misrepair was found to be approximately 10 - 15% (Lobrich et al. 2000); this rate increased to 50 - 60% at a radiation exposure of 80 Gy (Kuhne et al. 2000; Lobrich et al. 2000; Mcmahon et al. 2016). It is not known, however, how this rate of misrepair relates to chromosomal aberration frequency. Results from one study using a DNA repair inhibitor suggested that as adequate DNA repair declines, the chromosomal aberration frequency increases (Chernikova et al. 1999).  The time scale between inadequate repair and chromosomal aberration frequency has also not been well established.

Mutations, Increase (KE3)    --> Cell Proliferation, Increase (KE5)

Evidence for Quantitative Understanding of KER: Weak

 Quantitative understanding of the relationship between these two events has not been well established. There are, however, some studies that have examined how cellular proliferation changes over time in the presence of mutations. In cells harbouring mutations in critical genes, higher proliferation rates were evident by the fourth day in culture (Lang et al. 2004; Li and Xiong 2017) and higher rates of population doublings were evident by passage 7 (Li and Xiong 2017) relative to wild-type cells. DNA synthesis (which could be indicative of cellular proliferation) was higher in p53-/- cells than in wild-type cells for the first 6 days of culture, and increased to drastically higher levels in the knock-out cells until the end of the experiment at day 10 (Lang et al. 2004). In vivo, mice injected with oncogenically-transformed cells containing multiple mutations had detectable tumour growth by 10 - 12 days post-inoculation. These volumes continued increasing over the 40-day experiment (Sato et al. 2017).  

Chromosomal Aberrations, Increase (KE4)    --> Cell Proliferation, Increase (KE5)

Evidence for Quantitative Understanding of KER: Weak

Quantitative understanding of the relationship between these two events has not been well established. . Although studies that directly assessed the time scale between chromosomal aberrations and cell proliferation rates were not identified, differences in cellular proliferation rates for cells with different CA-related manipulations or treatments were evident within the first 3 days of culture (Stopper et al. 2003; Li et al. 2007; Soda et al. 2007; Irwin et al. 2013; Guarnerio et al. 2016).

Cell Proliferation, Increase (KE5)    --> Lung Cancer, Increase (AO)

Evidence for Quantitative Understanding of KER: Weak

Quantitative understanding of the relationship between these two events has not been well established. Human non-carcinogenic cells are thought to undergo 50 – 70 cell divisions before the telomeres can no longer support cell division (Panov 2005); this number would presumably be higher in cancer cells, but  quantitative data was not able to be identified. There are some studies available, however, that provide some details regarding the timing between these two events. In vitro experiments using lung cancer cell lines demonstrated that expression levels of key proteins involved in the regulation of the cell cycle and/or proliferation were modified by chemical inhibitors within the first 48 hours of treatment (Lv et al. 2012; Wanitchakool et al. 2012; Pal et al. 2013; Sun et al. 2016). In vivo studies using xenograft nude mice found that tumours were detected within two weeks of NSCLC-cell inoculation, and continued to grow over the experimental period (Pal et al. 2013; Warin et al. 2014; Sun et al. 2016; Tu et al. 2018). Differences in tumour growth rates between mice treated with an anti-cancer drug and those left untreated were also evident within 13 - 27 days (Pal et al. 2013; Sun et al. 2016; Tu et al. 2018), with significant differences in cell proliferation markers and tumour numbers or sizes at time of harvest (22 days - 27 weeks) (Kassie et al. 2008; Pal et al. 2013; Warin et al. 2014; Sun et al. 2016; Tu et al. 2018).

Double-Strand Breaks, Increase (KE1) --> Mutations, Increase (KE3)

Evidence for Quantitative Understanding of KER: Weak

There is overall limited quantitiative understanding of the relationship between DSBs and increased mutation rates. McMahon et al., 2016 compiled data from multiple studies spanning different human and mouse cell lines to model the IR dose-dependent increase in mutation rate. However, further quantitiative studies into this relationship are required to provide a better quantitiative understanding.

Double-Strand Breaks, Increase (KE1) --> Chromosomal Aberrations, Increase (KE4)

Evidence for Quantitative Understanding of KER: Weak

Similarly to the non-adjacent relationship above (KE1 -> KE4), there is overall limited quantitiative understanding of the relationship between DSBs and increased rates of chromosomal aberrations. McMahon et al., 2016 compiled data from multiple studies spanning different human and mouse cell lines to model the IR dose-dependent increase in the rate of chromosomal aberrations. However, further quantitiative studies into this relationship are required to provide a better quantitiative understanding.

Mutations, Increase (KE3)    --> Lung Cancer, Increase (AO)

Evidence for Quantitative Understanding of KER: Weak

Finding studies addressing the quantitative relationship between mutations and cancer directly was particularly challenging. However, many studies indicated that there was a positive, dose-dependent increase in mutations with increasing radiation dose (Suzuki and Hei 1996; Canova et al. 2002). A similar positive, dose-dependent relationship was found for the oncogenic transformations in cell and the radiation dose (Miller et al. 1995), and the incidence of lung cancer in rats and their cumulative radon exposure (Monchaux et al. 1994). Epidemiological studies examining lung cancer in radon-exposed uranium miners found a positive, linear relationship between lung cancer and cumulative radon exposure (Lubin et al. 1995; Ramkissoon et al. 2018). In terms of time-scale, mutations were evident in 2 weeks following irradiation (Hei et al. 1997), whereas oncogenic transformations took 7 weeks to develop following radiation exposure (Miller et al. 1999). In vivo models with injected tumour cells, inherent mutations, exposure to carcinogens, or Cre-induced mutations showed tumour growth months after exposure to the tumour-inducing insult (Hei et al. 1994; Fisher et al. 2001; Kasinski and Slack 2012; Fujimoto et al. 2017).

Chromosomal Aberrations, Increase (KE4)    --> Lung Cancer, Increase (AO)

Evidence for Quantitative Understanding of KER: Moderate

There is evidence of a positive, linear relationship between radiation dose and CAs (Nagasawa et al. 1990; A.L. Brooks et al. 1995; Khan et al. 1995; Yamada et al. 2002; Stevens et al. 2014), radiation dose and oncogenic transformations (Miller et al. 1996), as well as radon exposure and the risk of lung cancer mortality (Tirmarchel et al. 1993; Walsh et al. 2010). The latter relationship was found to be exponentially modified, however, by factors such as the age at median exposure, the time since median exposure, and the radon exposure rate (Walsh et al. 2010). Equations defining these relationships were derived in a number of different studies (Tirmarchel et al. 1993; A.L. Brooks et al. 1995; Khan et al. 1995; Miller et al. 1996; Girard et al. 2000; Yamada et al. 2002; Walsh et al. 2010; Stevens et al. 2014). In terms of time scale, micronuclei were documented in cells of the rodent lung as early as 0.2 days (Khan et al. 1995), and were found to persist for days to weeks (Khan et al. 1995; Deshpande et al. 1996; Werner et al. 2017). Oncogenic transformations, on the other hand, took weeks to develop (Robertson et al. 1983; Miller et al. 1996), while lung tumours took months or years to develop following radiation exposure (Tirmarchel et al. 1993; Yamada et al. 2017). Delivery of an agent carrying a cancer-related CA resulted in tumour growth within 21 - 31 days of its injection into mice (Pear et al. 1998; Kuramochi et al. 2001).

Direct Deposition of Energy (MIE) --> Lung Cancer, Increase (AO)

Evidence for Quantitative Understanding of KER: Moderate

Quantitative understanding has been well-established for this KER. According to current Canadian guidelines developed by Health Canada, annual residential radon levels should not exceed 200 Bq/m3. Similarly, the WHO recommends that the national annual residential radon levels not exceed 100 Bq/m3 where possible; if there are geographic or national constraints that make this target unachievable, the national standard should not be higher than 300 Bq/m3 (World Health Organization - Radon Guide 2009). Positive relationships between radon exposure and lung cancer have been established using in vitro models (Miller 1995), in vivo models(Monchaux et al. 1994) and results from human epidemiological studies (Lubin et al. 1995; Hazelton et al. 2001; Darby et al. 2005; Krewski et al. 2005; Krewski et al. 2006; Rodríguez-Martínez et al. 2018; Ramkissoon et al. 2018). Unsurprisingly, oncogenic transformation in cells were found weeks after radiation exposure (Miller et al. 1995), sizable tumours developed months after irradiation in mice (Hei et al. 1994) and lung cancer was found years after exposure in humans (Lubin et al. 1995; Darby et al. 2005; Torres-Durán et al. 2014; Rodríguez-Martínez et al. 2018; Ramkissoon et al. 2018).


Considerations for Potential Applications of the AOP (optional)

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At present the AOP framework is not readily used to support regulatory decision-making in radiation protection practices. The goal of developing this AOP is to bring attention to the framework to the radiation community as an effective means to organize knowledge,  identify gaps  and co-ordinate research.  We have used lung cancer as the case example due to its relevance to radon risk assessment and broadly because it can be represented as a simplified targeted path with a molecular initiating event that is specific to a radiation insult.  From this AOP, more complex networks can form which are relevant to both radiation and chemical exposure scenarios. Furthermore, as  mechanistic knowledge surrounding low dose radiation exposures becomes clear, this information can be incorporated into the AOP.  By developing this AOP, we have supported the necessary efforts highlighted by the international and national radiation protection agencies such as, the United Nations Scientific Committee on the Effects of Atomic Radiation, International Commission of Radiological Protection, International Dose Effect Alliance and the Electric Power Research Institute Radiation Program to consolidate and enhance the knowledge in understanding of low dose radiation exposures from the cellular to organelle levels within the biological system.


References

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Abe Y, Yoshida MA, Fujioka K, Kurosu Y, Ujiie R, Yanagi A, Tsuyama N, Miura T, Inaba T, Kamiya K, et al. 2018. Dose – response curves for analyzing of dicentric chromosomes and chromosome translocations following doses of 1000 mGy or less , based on irradiated peripheral blood samples from fi ve healthy individuals. 59(1):35–42. doi:10.1093/jrr/rrx052.

Adewoye AB, Lindsay SJ, Dubrova YE, Hurles ME. 2015. mutation induction in the mammalian germline. doi:10.1038/ncomms7684.

Al-Zoughool M, Krewski D. 2009. Health effects of radon: A review of the literature. Int J Radiat Biol. 85(1):57–69. doi:10.1080/09553000802635054.

Albertini RJ, Clark LS, Nicklas JA, Neill JPO, Hui E, Jostes R, Albertini RJ, Clark LS, Nicklas JA, Neill JPO, et al. 1997. Radiation Quality Affects the Efficiency of Induction and the Molecular Spectrum of HPRT Mutations in Human T Cells. 148(5).

Albertson DG, Collins C, Mccormick F, Gray JW. 2003. Chromosome aberrations in solid tumors. 34(4):369–376.

Alexandrov LB, Nik-zainal S, Wedge DC, Aparicio SAJR, Behjati S, Biankin A V, Bignell GR, Bolli. 2013. Signatures of mutational processes in human cancer. doi:10.1038/nature12477.

Ali HEA, Barber RC, Dubrova YE. 2012. Mutation Research / Fundamental and Molecular Mechanisms of Mutagenesis The effects of maternal irradiation during adulthood on mutation induction and transgenerational instability in mice. 732:21–25. doi:10.1016/j.mrfmmm.2012.01.003.
Amundson SA, Chen DJ.1996.Ionizing radiation-induced mutation of human cells with different DNA repair capacities. Advances in Space Research, 18(1-2):119-126. doi: doi:10.1016/0273-1177(95)00798-J

Antonelli AF, Campa A, Esposito G, Giardullo P, Belli M, Dini V, Meschini S, Simone G, Sorrentino E, Gerardi S, et al. 2015. Induction and Repair of DNA DSB as Revealed by H2AX Phosphorylation Foci in Human Fibroblasts Exposed to Low- and High-LET Radiation : Relationship with Early and Delayed Reproductive Cell Death Induction and Repair of DNA DSB as Revealed by H2AX Phosphor. doi:10.1667/RR13855.1.

Arlt MF, Rajendran S, Birkeland SR, Wilson TE, Glover TW. 2014. NIH Public Access. 55(2):103–113. doi:10.1002/em.21840.

Asaithamby A, Chen DJ. 2009. Cellular responses to DNA double-strand breaks after low-dose c -irradiation. 37(12):3912–3923. doi:10.1093/nar/gkp237.

Axelson O. 1995. Cancer risks from exposure to radon in homes. Environ Health 1995. (8):37–43.

Balajee AS, Bertucci A, Taveras M, Brenner DJ. 2014. Multicolour FISH analysis of ionising radiation induced micronucleus formation in human lymphocytes. 29(6):447–455. doi:10.1093/mutage/geu041.

Barber RC, Hardwick RJ, Shanks ME, Glen CD, Mughal SK, Voutounou M, Dubrova YE. 2009. Mutation Research / Fundamental and Molecular Mechanisms of Mutagenesis The effects of in utero irradiation on mutation induction and transgenerational instability in mice. 664:6–12. doi:10.1016/j.mrfmmm.2009.01.011.

Beir V. 1999b. The Mechanistic Basis of Radon-Induced Lung Cancer. https://www.ncbi.nlm.nih.gov/books/NBK233261/.

Barron CC, Moore J, Tsakiridis T, Pickering G, Tsiani E. 2014. Inhibition of human lung cancer cell proliferation and survival by wine. Cancer Cell Int. 14(1):1–13. doi:10.1186/1475-2867-14-6.

Bartova E, Kozubek S, Kozubek M, Jirsova P, Lukasova E, Skalnikova M, Buchnickova K. 2000. The influence of the cell cycle , differentiation and irradiation on the nuclear location of the abl , bcr and c - myc genes in human leukemic cells. Leukemia Research. 24(3):233-41.doi: 10.1016/S0145-2126(99)00174-5.

Basheerudeen SS, Kanagaraj K, Jose MT, Ozhimuthu A, Paneerselvam S, Pattan S, Joseph S, Raavi V. 2017. Entrance surface dose and induced DNA damage in blood lymphocytes of patients exposed to low-dose and low-dose-rate X-irradiation during diagnostic and therapeutic interventional radiology procedures. Mutat Res Gen Tox En. 818(April):1–6. doi:10.1016/j.mrgentox.2017.04.001.

Beels L, Bacher K, Wolf D De, Werbrouck J, Thierens H. 2009. -H2AX Foci as a Biomarker for Patient X-Ray Exposure in Pediatric Cardiac Catheterization Are We Underestimating Radiation Risks ? :1903–1909. doi:10.1161/circulationa.109.880385.

Behjati S, Gundem G, Wedge DC, Roberts ND, Tarpey PS, Cooke SL, Loo P Van, Alexandrov LB, Ramakrishna M, Davies H, et al. 2016. in second malignancies. :1–8. doi:10.1038/ncomms12605.

Bertram JS. 2001. The molecular biology of cancer. Mol Aspects Med. 21:166–223. doi:10.1016/S0098-2997(00)00007-8.

Bétermier M, Bertrand P, Lopez BS. 2014. Is Non-Homologous End-Joining Really an Inherently Error-Prone Process? PLoS Genet. 10(1). doi:10.1371/journal.pgen.1004086.

Bignold LP. 2009. Mutation Research / Reviews in Mutation Research Mechanisms of clastogen-induced chromosomal aberrations : A critical review and description of a model based on failures of tethering of DNA strand ends to strand-breaking enzymes. 681:271–298. doi:10.1016/j.mrrev.2008.11.004.

Boffetta P, Hel O Van Der, Norppa H, Fabianova E, Fucic A, Gundy S, Lazutka J, Cebulska-wasilewska A, Puskailerova D, Znaor A, et al. 2007. Original Contribution Chromosomal Aberrations and Cancer Risk : Results of a Cohort Study from Central Europe. 165(1):36–43. doi:10.1093/aje/kwj367.

Bolsunovsky A, Frolova T, Dementyev D, Sinitsyna O. 2016. Ecotoxicology and Environmental Safety Low doses of gamma-radiation induce SOS response and increase mutation frequency in Escherichia coli and Salmonella typhimurium cells. Ecotoxicol Environ Saf. 134:233–238. doi:10.1016/j.ecoenv.2016.09.009.

Bonassi S, Hagmar L, Stro U, Montagud AH, Tinnerberg H, Forni A, Wanders S, Wilhardt P, Hansteen I, Knudsen LE, et al. 2000. Chromosomal Aberrations in Lymphocytes Predict Human Cancer Independently of Exposure to Carcinogens 1. Cancer Research 60(6):1619–1625.

Bonassi S, Norppa H, Ceppi M, Vermeulen R, Znaor A, Fabianova E, Gundy S, Hansteen I. 2008. Chromosomal aberration frequency in lymphocytes predicts the risk of cancer : results from a pooled cohort study of 22 358 subjects in 11 countries. 29(6):1178–1183. doi:10.1093/carcin/bgn075.

Bracalente C, Ibanez IL, Molinari B, Palmieri M, Kreiner A, Valda A, Davidson J, Duran H. 2013. Induction and Persistence of Large g H2AX Foci by High Linear Energy Transfer Radiation in DNA-Dependent protein kinase e Deficient Cells. 87(4). doi:10.1016/j.ijrobp.2013.07.014.

Brooks AAL, Miick R, Buschbom RL, Murphy MK, Khan MA, Brooks AL, Miick R, Buschbom RL, Murphy MK, Khan MA. 1995. The Role of Dose Rate in the Induction of Micronuclei in Deep-Lung Fibroblasts In Vivo after Exposure to Cobalt-60 Gamma Rays The Role of Dose Rate in the Induction of Micronuclei in Deep-L Fibroblasts In Vivo after Exposure to Cobalt-60 Gamma Ray. Radiation Research. 144(1):114-8. doi:10.2307/3579244.

Burr KL, Duyn-goedhart A Van, Hickenbotham P, Monger K, Buul PPW Van, Dubrova YE. 2007. The effects of MSH2 deficiency on spontaneous and radiation-induced mutation rates in the mouse germline. 617:147–151. doi:10.1016/j.mrfmmm.2007.01.010.

Canova S, Perin M, Mognato M, Favaretto S, Cherubini R, Celotti L. 2002. Minisatellite and HPRT Mutations in V79 And Human Cells Irradiated with Gamma Rays. 99:207–209. doi:10.1093/oxfordjournals.rpd.a006763.

Chang HHY, Pannunzio NR, Adachi N, Lieber MR. 2017. Non-homologous DNA end joining and alternative pathways to double ‑ strand break repair. Nat Publ Gr. 18(8):495–506. doi:10.1038/nrm.2017.48.

Charlton DE, Nikjoo H, Humm JL. 1989. Calculation of initial yields of single- and double-strand breaks in cell nuclei from electrons , protons and alpha particles.

Cheki M, Shirazi A, Mahmoudzadeh A, Tavakkoly J. 2016. Mutation Research / Genetic Toxicology and Environmental Mutagenesis The radioprotective effect of metformin against cytotoxicity and genotoxicity induced by ionizing radiation in cultured human blood lymphocytes. Mutat Res - Genet Toxicol Environ Mutagen. 809:24–32. doi:10.1016/j.mrgentox.2016.09.001.

Chernikova ASB, Wells RL, Elkind MM, Chernikova SB, Wells RL, Elkind MM. 1999. Wortmannin Sensitizes Mammalian Cells to Radiation by Inhibiting the DNA-Dependent Protein Kinase-Mediated Rejoining of Double-Strand Breaks Linked references are available on JSTOR for this article : Wortmannin Sensitizes Mammalian Cells to Radiation by. 151(2):159–166.

Christensen DM. 2014. Management of Ionizing Radiation Injuries and Illnesses, Part 3: Radiobiology and Health Effects of Ionizing Radiation. 114(7):556–565. doi:10.7556/jaoa.2014.109.
Cornforth M, Bedford J. 1985. On the Nature of a Defect in Cells from Individuals with Ataxia-Telangiectasia. (16):1589–1592. doi:10.1126/science.3975628.

Cortot AB, Younes ÃM, Martel-planche ÃG, Guibert B, Isaac S, Souquet P, Commo F, Girard P, Fouret P, Brambilla E, et al. 2014. Mutation of TP53 and Alteration of p14 arf Expression in EGFR- and KRAS -Mutated Lung Adenocarcinomas. (March):124–130. doi:10.1016/j.cllc.2013.08.003.

Danford N. 2012. The Interpretation and Analysis of Cytogenetic Data. 817. doi:10.1007/978-1-61779-421-6.

Darby S, Hill D, Auvinen A, Barros-Dios JM, Baysson H, Bochicchio F, Deo H, Falk R, Forastiere F, Hakama M, et al. 2005. Radon in homes and risk of lung cancer: Collaborative analysis of individual data from 13 European case-control studies. Br Med J. 330(7485):223–226. doi:10.1136/bmj.38308.477650.63.

Deshpande AA, Goodwin EH, Bailey SM, Marrone BL, Lehnert BE, Mar N, Deshpande A, Goodwin EH, Bailey SM, Marrone BL, et al. 1996. Alpha-Particle-Induced Sister Chromatid Exchange in Normal Human Lung Fibroblasts . 145(3):260–267.

Desouky O, Ding N, Zhou G. 2015. ScienceDirect Targeted and non-targeted effects of ionizing radiation. J Radiat Res Appl Sci. 8(2):247–254. doi:10.1016/j.jrras.2015.03.003.
Dikomey E, Brammer I. 2000. Relationship between cellular radiosensitivity and non-repaired double-strand breaks studied for di Ú erent growth states , dose rates and plating conditions in a normal human broblast line. 76(6). doi:10.1080/09553000050028922.

Dong J, Zhang T, Ren Y, Wang Z, Ling CC, He F, Li GC, Wang C, Wen B. 2017. Inhibiting DNA-PKcs in a non-homologous end-joining pathway in response to DNA double-strand breaks. Oncotarget. 8(14):22662–22673. doi:10.18632/oncotarget.15153.

Duan W, Gao ÆL, Jin ÆD, Villalona-calero. 2008. Lung specific expression of a human mutant p53 affects cell proliferation in transgenic mice. :355–366. doi:10.1007/s11248.

Dubrova YE, Grant G, Chumak AA, Stezhka VA, Karakasian AN. 2002. Elevated Minisatellite Mutation Rate in the Post-Chernobyl Families from Ukraine. 32:801–809. doi: 10.1086/342729.

Dubrova YE, Plumb M, Brown J, Boulton E, Goodhead D, Jeffreys AJ. 2000. Induction of minisatellite mutations in the mouse germline by low-dose chronic exposure to  -radiation and fission neutrons. 453:17–24.

Dubrova YE, Plumb M, Brown J, Fennelly J, Bois P, Goodhead D, Jeffreys AJ. 1998. Stage specificity , dose response , and doubling dose for mouse minisatellite germ-line mutation induced by acute radiation. 95(May):6251–6255. doi: 10.1073/pnas.95.11.6251.

Dubrova YE, Plumb MA. 2002. Ionising radiation and mutation induction at mouse minisatellite loci The story of the two generations. 499:143–150. doi: 10.1016/s0027-5107(01)00284-6.


El-zein RA, Kyle A, Santee J, Yu R. 2017. Identification of Small and Non-Small Cell Lung Cancer Markers in Peripheral Blood Using Cytokinesis-Blocked Micronucleus and Spectral Karyotyping Assays. 77030:122–131. doi:10.1159/000479809.

El-zein RA, Lopez MS, Jr AMDA, Liu M, Munden RF, Christiani D, Su L, Tejera-alveraz P, Zhai R, Spitz MR, et al. 2014. The Cytokinesis-Blocked Micronucleus Assay as a Strong Predictor of Lung Cancer : Extension of a Lung Cancer Risk Prediction Model. 23(November):2462–2471. doi:10.1158/1055-9965.EPI-14-0462.

Eymin B, Gazzeri S. 2009. Role of cell cycle regulators in lung carcinogenesis. 4(1):114–123. doi: 10.4161/cam.4.1.10977.

Feldmann E, Schmiemann V, Goedecke W, Reichenberger S, Pfeiffer P, Essen U, Essen D-. 2000. DNA double-strand break repair in cell-free extracts from Ku80-deficient cells : implications for Ku serving as an alignment factor in non-homologous DNA end joining. 28(13):2585–2596. doi: 10.1093/nar/28.13.2585.

Fenech M, Natarajan AT. 2011. Molecular mechanisms of micronucleus , nucleoplasmic bridge and nuclear bud formation in mammalian and human cells. 26(1):125–132. doi:10.1093/mutage/geq052.

Ferguson DO, Alt FW. 2001. DNA double strand break repair and chromosomal translocation : Lessons from animal models. :5572–5579. doi: 10.1038/sj.onc.1204767.

Fisher GH, Wellen SL, Klimstra D, Lenczowski JM, Tichelaar JW, Lizak MJ, Whitsett JA, Koretsky A, Varmus HE. 2001. Induction and apoptotic regression of lung adenocarcinomas by regulation of a K-Ras transgene in the presence and absence of tumor suppressor genes. :3249–3262. doi:10.1101/gad.947701.NSCLCs.

Flegal M, Blimkie MS, Wyatt H, Bugden M, Surette J, Klokov D. 2015. Measuring DNA Damage and Repair in Mouse Splenocytes After Chronic In Vivo Exposure to Very Low Doses of Beta- and Gamma-Radiation. (July):1–9. doi:10.3791/52912.

Fujimoto J, Nunomura-nakamura S, Liu Y, Lang W, Mcdowell T, Jakubek Y, Ezzeddine D, Ochieng JK, Petersen J, Davies G, et al. 2017. Development of Kras mutant lung adenocarcinoma in mice with knockout of the airway lineage-specific gene Gprc5a. 141(8):1589–1599. doi:10.1002/ijc.30851.Development.

Geng C, Kaochar S, Li M, Rajapakshe K, Fiskus W, Dong J, Foley C, Dong B, Zhang L, Kwon O, et al. 2017. ORIGINAL ARTICLE SPOP regulates prostate epithelial cell proliferation and promotes ubiquitination and turnover of c-MYC oncoprotein. (January):4767–4777. doi:10.1038/onc.2017.80.

Van Gent DC, Hoeijmakers JHJ, Kanaar R. 2001. Chromosomal Stability And The DNA Double-Stranded Break Connection. 2(March):196–206. doi: 10.1038/35056049.

George AKA, Hada M, Jackson LJ, Elliott T, Kawata T, George KA, Hada M, Jackson LJ, Elliott T, Kawata T, et al. 2009. Dose Response of γ Rays and Iron Nuclei for Induction of Chromosomal Aberrations in Normal and Repair-Deficient Cell Lines Dose Response of c Rays and Iron Nuclei for Induction of Chromosomal Aberrations in Normal and Repair-Deficient Cell Lines. 171(6):752–763. doi: 10.1667/RR1680.1


Ghazavi F, Lammens T, Roy N Van, Poppe B, Speleman F, Benoit Y, Vlierberghe P Van, Moerloose B De. 2015. Molecular basis and clinical significance of genetic aberrations in B-cell precursor acute lymphoblastic leukemia. Exp Hematol. 43(8):640–653. doi:10.1016/j.exphem.2015.05.015.

Girard L, Zo S, Virmani AK, Gazdar AF, Minna JD. 2000. Genome-wide Allelotyping of Lung Cancer Identifies New Regions of Allelic Loss , Differences between Small Cell Lung Cancer and Non-Small Cell Lung Cancer , and Loci Clustering 1. :4894–4906.

Goodhead DT. 2006. Energy deposition stochastics and track structure: What about the target? Radiat Prot Dosimetry. 122(1–4):3–15. doi:10.1093/rpd/ncl498.

Gossen JA, Martus H-J, Wei Y, Vijg J. 1995. Spontaneous and  X-ray-induced deletion mutations in a LacZ plasmid-based transgenic mouse model. 331:89–97. doi: 10.1016/0027-5107(95)00055-n.

Gronroos E. 2018. Tolerance of Chromosomal Instability in Cancer : Mechanisms and Therapeutic Opportunities. :0–1. doi:10.1158/0008-5472.CAN-18-1958.

Grudzenski S, Raths A, Conrad S, Rübe CE, Löbrich M. 2010. Inducible response required for repair of low-dose radiation damage in human fi broblasts. doi:10.1073/pnas.1002213107.

Guarnerio J, Bezzi M, Jeong JC, Tay Y, Beck AH, Pandolfi PP, Guarnerio J, Bezzi M, Jeong JC, Paffenholz S V, et al. 2016. Oncogenic Role of Fusion-circRNAs Derived from Article Oncogenic Role of Fusion-circRNAs Derived from Cancer-Associated Chromosomal Translocations. Cell. 165(2):289–302. doi:10.1016/j.cell.2016.03.020.

Hada M, Georgakilas AG. 2008. Formation of Clustered DNA Damage after High-LET Irradiation : A Review. 49(3):203–210. doi:10.1269/jrr.07123.

Hagmar L, Stro U, Bonassi S, Hansteen I, Knudsen LE, Lindholm C. 2004. Impact of Types of Lymphocyte Chromosomal Aberrations on Human Cancer Risk : Results from Nordic and Italian Cohorts. :2258–2263. doi: 10.1158/0008-5472.CAN-03-3360.

Han L, Zhao F, Sun Q, Wang P, Wang X, Guo F, Fu B, Lü Y. 2014. Mutation Research / Genetic Toxicology and Environmental Mutagenesis Cytogenetic analysis of peripheral blood lymphocytes , many years after exposure of workers to low-dose ionizing radiation. 771:1–5.

Hanahan D, Weinberg RA. 2011. Review Hallmarks of Cancer : The Next Generation. Cell. 144(5):646–674. doi:10.1016/j.cell.2011.02.013.

Hazelton WD, Luebeck EG, Heidenreich WF, Moolgavkar SH. 2001. Analysis of a Historical Cohort of Chinese Tin Miners with Arsenic, Radon, Cigarette Smoke, and Pipe Smoke Exposures Using the Biologically Based Two-Stage Clonal Expansion Model. Radiat Res. 156(1):78–94. doi:10.1667/0033-7587(2001)156[0078:aoahco]2.0.co;2.

Hei TK, Piao CQ, Willey JC, Thomas S, Hal EJ, Ny R. 1994. Malignant transformation of human bronchial epithelial cells by radon-simulated or-particles. 15(3):431–437. doi: 10.1093/carcin/15.3.431.


Hei TK, Wu L-J, Liu S-X, Vannais D, Waldren C, Randers-pehrson G. 1997. Mutagenic effects of a single and an exact number of ␣ particles in mammalian cells. 94(April):3765–3770.
Heng HHQ, Bremer SW, Stevens J, Ye KJ, Miller F, Liu G, Ye CJ. 2006. Cancer Progression by Non-Clonal Chromosome Aberrations. 1435:1424–1435. doi:10.1002/jcb.20964.

Heng HHQ, Stevens JB, Liu GOU, Bremer SW, Ye KJ, Reddy P, Wu GENS, Wang YA, Tainsky MA, Ye CJ, et al. 2006. Stochastic Cancer Progression Driven by Non-Clonal Chromosome Aberrations. 472:461–472. doi:10.1002/jcp.

Heterodimer K, Gaymes TJ, Mufti GJ, Rassool F V. 2002. Myeloid Leukemias Have Increased Activity of the Nonhomologous End-Joining Pathway and Concomitant DNA Misrepair that Is Dependent on the  the Ku70/86 heterodimer.:2791–2797.

Hoeijmakers JHJ. 2001a. Genome Maintenance for Preventing Cancer. DNA Repair (Amst). 411:366–374. doi:10.1038/35077232.

Hundley JE, Koester SK, Troyer DA, Hilsenbeck SG, Subler MA, Windle JJ. 1997. Increased Tumor Proliferation and Genomic Instability without Decreased Apoptosis in MMTV- ras Mice Deficient in p53. Mol Cell Biol. 17(2):723–731. doi:10.1128/MCB.17.2.723.

Irwin ME, Nelson LD, Farrill JMS, Knouse PD, Miller CP, Palla SL, Siwak DR, Mills GB, Estrov Z, Li S, et al. 2013. Small Molecule ErbB Inhibitors Decrease Proliferative Signaling and Promote Apoptosis in Philadelphia Chromosome – Positive Acute Lymphoblastic Leukemia. 8(8):1–10. doi:10.1371/journal.pone.0070608.

Iwakuma T, Lozano G. 2007. Crippling p53 activities via knock-in mutations in mouse models. :2177–2184. doi:10.1038/sj.onc.1210278.

Jang M, Han E, D JKLM, Cho KH, Ph D, Shin HB, Ph D, Lee YK, Ph D. 2019. Dose Estimation Curves Following In Vitro X-ray Irradiation Using Blood From Four Healthy Korean Individuals. :91–95. doi: 10.3343/alm.2019.39.1.91.

Jeggo PA, Markus L. 2015. How cancer cells hijack DNA double-strand break repair pathways to gain genomic instability. :1–11. doi:10.1042/BJ20150582.

Jeggo PA, Markus L. 2015. How cancer cells hijack DNA double-strand break repair pathways to gain genomic instability. :1–11. doi:10.1042/BJ20150582.

Jia P, Pao W, Zhao Z. 2014. Patterns and processes of somatic mutations in nine major cancers. BMC Med Genomics. 7(1):1–11. doi:10.1186/1755-8794-7-11.

Jia Y, Juarez J, Li J, Manuia M, Niederst MJ, Tompkins C, Timple N, Vaillancourt M, Pferdekamper AC, Lockerman EL, et al. 2016. EGF816 Exerts Anticancer Effects in Non – Small Cell Lung Cancer by Irreversibly and Selectively Targeting Primary and Acquired Activating Mutations in the EGF Receptor. 1686(15):1591–1603. doi:10.1158/0008-5472.CAN-15-2581.

Joiner M. 2009. Basic Clinical Radiobiology Edited by. [1] PJ Sadler, Next-Generation Met Anticancer Complexes Multitargeting via Redox Modul Inorg Chem 52 21.:375. doi:10.1201/b13224.

Jorge S-G, Breña-Valle M, Aguilar-Moreno M, Balcázar M. 2012. Evidence of DNA double strand breaks formation in Escherichia coli bacteria exposed to alpha particles of different LET assessed by the SOS response. Appl Radiat Isot. 71(SUPPL.):66–70. doi:10.1016/j.apradiso.2012.05.007.

Jostes RF. 1996. Genetic , cytogenetic , and carcinogenic effects of radon : a review. Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis. 340(2-3):125-39. doi: 10.1016/S0165-1110(96)90044-5.

Kang ZJ, Liu YF, Xu LZ, Long ZJ, Huang D, Yang Y, Liu B, Feng JX. 2016. The Philadelphia chromosome in leukemogenesis. Chin J Cancer.:1–15. doi:10.1186/s40880-016-0108-0.

Karanjawala ZE, Grawunder U, Hsieh C, Lieber MR. The nonhomologous DNA end joining pathway is important for chromosome stability in primary fibroblasts. Current Biology 9(24):1501-4. doi: 10.1016/S0960-9822(00)80123-2.

Kasinski AL, Slack FJ. 2012. miRNA-34 Prevents Cancer Initiation and Progression in a Therapeutically Resistant K-ras and p53-Induced Mouse Model of Lung Adenocarcinoma. 72(11):5576–5588. doi:10.1158/0008-5472.

Kassie F, Matise L, Negia M, Upadhyaya P, Hecht SS. 2008. NIH Public Access. 1(7):1–16. doi:10.1158/1940-6207.

Kendall GM, Smith TJ. 2002a. Doses to organs and tissues from radon and its decay products. J Radiol Prot. 22(4):389–406. doi: 10.1088/0952-4746/25/3/002.

Khan MA, Cross FT, Buschbom RL, Brooks AL. 1995. Inhaled radon-induced genotoxicity in Wistar rat , Syrian hamster , and Chinese hamster deep-lung fibroblasts in vivo. 334:131–137.
Khanna KK, Jackson SP. 2001. DNA double-strand breaks : signaling , repair and the cancer connection. 27(march):247–254.

Kim HR, Shim HS, Chung J, Lee YJ, Hong YK. 2012. Distinct Clinical Features and Outcomes in Never-Smokers With Nonsmall Cell Lung Cancer Who Harbor EGFR or KRAS Mutations or ALK Rearrangement. :729–739. doi:10.1002/cncr.26311.

Kim MP, Lozano G. 2018. Mutant p53 partners in crime. Nat Publ Gr. 25(1):161–168. doi:10.1038/cdd.2017.185.

Krewski D, Lubin JH, Zielinski JM, Alavanja M, Catalan VS, Field RW, Klotz JB, L.tourneau EG, Lynch CF, Lyon JI, et al. 2005. Residential Radon and Risk of Lung Cancer. Epidemiology. 16(2):137–145. doi:10.1097/01.ede.0000152522.80261.e3.

Krewski D, Lubin JH, Zielinski JM, Alavanja M, Catalan VS, Field RW, Klotz JB, Létourneau EG, Lynch CF, Lyon JL, et al. 2006. A combined analysis of north American case-control studies of residential radon and lung cancer. J Toxicol Environ Heal - Part A. 69(7–8):533–597. doi:10.1080/15287390500260945.

Kuefner MA, Grudzenski S, Schwab SA, Wiederseiner M. 2009. DNA Double-Strand Breaks and Their Repair in Blood Lymphocytes of Patients Undergoing Angiographic Procedures. Investigative radiology. 44(8):440-6. doi:10.1097/RLI.0b013e3181a654a5.

Kuefner MA, Michael K, Grazia M, Larissa A, Uder M. 2015. Chemoprevention of Radiation-Induced DNA Double-Strand Breaks with Antioxidants. :1–6. doi:10.1007/s40134-014-0081-9.

Kuhne M, Rothkamm K, Lobrich M. 2000. No dose-dependence of DNA double-strand break misrejoining following a -particle irradiation. Genes Chromosomes Cancer.27(1):59-68.

Kuhne M, Urban G, Lo M. 2005. DNA Double-Strand Break Misrejoining after Exposure of Primary Human Fibroblasts to C K Characteristic X Rays , 29 kVp X Rays and Co g Rays. 676:669–676. doi: 10.1667/RR3461.1

Kuramochi M, Fukuhara H, Nobukuni T, Kanbe T, Maruyama T, Ghosh HP, Pletcher M, Isomura M, Onizuka M, Kitamura T, et al. 2001. TSLC1 is a tumor-suppressor gene in human non-small- cell lung cancer. 27(april):427–430. doi: 10.1038/86934.

Lang GA, Iwakuma T, Suh Y, Liu G, Rao VA, Parant JM, Valentin-vega YA, Terzian T, Caldwell LC, Strong LC, et al. 2004. Gain of Function of a p53 Hot Spot Mutation in a Mouse Model of Li-Fraumeni Syndrome. cell Press. 119(6):861–872. doi:10.1016/j.cell.2004.11.006.

Lara CM De, Hill MA, Jenner TJ, Papworth D. 2001. Dependence of the Yield of DNA Double-Strand Breaks in Chinese Hamster V79-4 Cells on the Photon Energy of Ultrasoft X Rays. 448:440–448. doi: 10.1667/0033-7587(2001)155[0440:DOTYOD]2.0.CO;2.

Larsen JE, Minna J. 2011. Molecular Biology of Lung Cancer : Clinical Implications. 32(4):703–740. doi:10.1016/j.ccm.2011.08.003.

Leibowitz ML, Zhang C, Pellman D. 2015. Chromothripsis : A New Mechanism for Rapid Karyotype Evolution. doi:10.1146/annurev-genet-120213-092228.

Lepage CC, Morden CR, Palmer MCL, Nachtigal MW, Mcmanus KJ. 2019. Detecting Chromosome Instability in Cancer : Approaches to Resolve Cell-to-Cell Heterogeneity. :1–20. doi:10.3390/cancers11020226.

Levine MS, Holland AJ. 2018. The impact of mitotic errors on cell proliferation and tumorigenesis. :620–638. doi:10.1101/gad.314351.118.620.

Li H, Ma X, Wang J, Koontz J, Nucci M, Sklar J. 2007. Effects of rearrangement and allelic exclusion of JJAZ1 / SUZ12 on cell proliferation and survival. 104(50):20001–20006. doi: 10.1073/pnas.0709986104.

Li Z, Xiong Y. 2017. Cytoplasmic E3 ubiquitin ligase CUL9 controls cell proliferation , senescence , apoptosis and genome integrity through p53. (November 2016):5212–5218. doi:10.1038/onc.2017.141.

Lieber MR, Gu J, Lu H, Shimazaki N, Tsai AG. 2010. Nonhomologous DNA End Joining (NHEJ) and Chromosomal Translocations in Humans. doi:10.1007/978-90-481-3471-7.

Lieber MR, Ma Y, Pannicke U, Schwarz K. 2003. Mechanism and regulation of human non-homologous DNA end-joining. Nat Rev Mol Cell Biol. 4(9):712–720. doi:10.1038/nrm1202.

Lim EH, Zhang S-L, Li J-L, Yap W-S, Howe T-C, Tan B-P, Lee Y-S, Wong D, Khoo K-L, Seto K-Y, et al. 2009. Using Whole Genome Amplification ( WGA ) of Low-Volume Biopsies to Assess the Prognostic Role of EGFR , KRAS , p53 ,. J Thorac Oncol. 4(1):12–21. doi:10.1097/JTO.0b013e3181913e28.

Lin Y, Nagasawa H, Little JB, Kato TA, Shih H, Xie X, Jr PFW, Brogan JR, Kurimasa A, Chen DJ, et al. 2014. Differential Radiosensitivity Phenotypes of DNA-PKcs Mutations Affecting NHEJ and HRR Systems following Irradiation with Gamma-Rays or Very Low Fluences of Alpha Particles. 9(4):2–11. doi:10.1371/journal.pone.0093579.

Liu M, Cai X, Yu W, Lv C, Fu X. 2015. Clinical significance of age at diagnosis among young non-small cell lung cancer patients under 40 years old: a population-based study. Oncotarget. 6(42). doi:10.18632/oncotarget.5524.

Liu X, Wang J, Chen L. 2013. Whole-exome sequencing reveals recurrent somatic mutation networks in cancer. Cancer Lett. 340(2):270–276. doi:10.1016/j.canlet.2012.11.002.

Lloyd SM, Lopez M, El-zein R. 2013. Cytokinesis-Blocked Micronucleus Cytome Assay and Spectral Karyotyping as Methods for Identifying Chromosome Damage in a Lung Cancer Case-Control Population. 707(September 2012):694–707. doi:10.1002/gcc.

Löbrich M, Cooper PK, Rydberg Björn, Lobrich M, Rydberg Bjorn. 1998. Joining of Correct and Incorrect DNA Ends at Double-Strand Breaks Produced by High-Linear Energy Transfer Radiation in Human Fibroblasts. Radiat Res. 150(6):619. doi:10.2307/3579884.

Lobrich M, Kuhne M, Wetzel J, Rothkamm K. 2000. Joining of Correct and Incorrect DNA Double-Strand Break Ends in Normal Human and Ataxia Telangiectasia Fibroblasts. 68:59–68.

Lomax ME, Folkes LK, Neill PO. 2013. Biological Consequences of Radiation-induced DNA Damage : Relevance to Radiotherapy Statement of Search Strategies Used and Sources of Information Why Radiation Damage is More Effective than Endogenous Damage at Killing Cells Ionising Radiation-induced Do. 25:578–585. doi:10.1016/j.clon.2013.06.007.

López-lázaro M. 2018. Critical Reviews in Oncology / Hematology The stem cell division theory of cancer. 123(July 2017):95–113. doi:10.1016/j.critrevonc.2018.01.010.

Lubin JH, Boice JD, Edling C, Richard W, Howe GR, Kunz E, Kusiak RA, Morrison I, Radford EP, Samet JM, et al. 1995. Lung Cancer in Radon-Exposed Miners and Estimation of Risk From Indoor Exposure.87(11) :817–827. doi: 10.1093/jnci/87.11.817.

Luo P, Wang Q, Ye Y, Zhang J, Lu D, Cheng L, Zhou H, Xie M, Wang B. 2019. MiR-223-3p functions as a tumor suppressor in lung squamous cell carcinoma by miR-223-3p-mutant p53 regulatory feedback loop. 1:1–12. doi: 10.1186/s13046-019-1079-1

Lv T, Yuan D, Miao X, Lv Y, Zhan P, Shen X, Song Y. 2012. Over-Expression of LSD1 Promotes Proliferation , Migration and Invasion in Non-Small Cell Lung Cancer. 7(4):1–8. doi:10.1371/journal.pone.0035065.

M Jarvis E, Kirk J, L Clarke C. 1998. Loss of Nuclear BRCA1 Expression in Breast Cancers Is Associated with a Highly Proliferative Tumor Phenotype.

Maffei F, Angelini S, Cantelli G, Violante FS, Lodi V, Mattioli S, Hrelia P. 2004. Spectrum of chromosomal aberrations in peripheral lymphocytes of hospital workers occupationally exposed to low doses of ionizing radiation. 547:91–99. doi:10.1016/j.mrfmmm.2003.12.003.

Maier P, Hartmann L, Wenz F, Herskind C. 2016. Cellular Pathways in Response to Ionizing Radiation and Their Targetability for Tumor Radiosensitization. doi:10.3390/ijms17010102.
Malu S, Malshetty V, Francis D, Cortes P. 2012. Role of non-homologous end joining in V(D)J recombination. Immunol Res. 54(1–3):233–246. doi:10.1007/s12026-012-8329-z.

Mao X, Boyd LK, Yáñez-muñoz RJ, Chaplin T, Xue L, Lin D, Berney DM, Young BD, Lu Y. 2011. Chromosome rearrangement associated inactivation of tumour suppressor genes in prostate cancer. 1(5):604–617.

Masumura K, Kuniya K, Kurobe T, Fukuoka M, Yatagai F, Nohmi T. 2002. Heavy-Ion-Induced Mutations in the gpt Delta Transgenic Mouse : Comparison of Mutation Spectra Induced by Heavy-Ion , X-Ray , and - Y-Ray Radiation. 215(June):207–215. doi:10.1002/em.10108.

Matuo Y, Izumi Y, Furusawa Y, Shimizu K. 2018. Mutat Res Fund Mol Mech Mutagen Biological e ff ects of carbon ion beams with various LETs on budding yeast Saccharomyces cerevisiae. Mutat Res Fund Mol Mech Mutagen. 810(November 2017):45–51. doi:10.1016/j.mrfmmm.2017.10.003.

Mcmahon SJ, Schuemann J, Paganetti H, Prise KM. 2016. Mechanistic Modelling of DNA Repair and Cellular Survival Following Radiation-Induced DNA Damage. Nat Publ Gr.(April):1–14. doi:10.1038/srep33290.

Meenakshi C, Mohankumar MN. 2013. Mutation Research / Genetic Toxicology and Environmental Mutagenesis Synergistic effect of radon in blood cells of smokers – An in vitro study. 757:79–82. doi: 10.1016/j.mrgentox.2015.06.008.

Meenakshi C, Sivasubramanian K, Venkatraman B. 2017. Mutation Research / Genetic Toxicology and Environmental Mutagenesis Nucleoplasmic bridges as a biomarker of DNA damage exposed to radon. Mutat Res - Genet Toxicol Environ Mutagen. 814:22–28. doi:10.1016/j.mrgentox.2016.12.004.

Mehta A, Haber JE. 2014. Sources of DNA Double-Strand Breaks and Models of Rec. Cold Spring Harb Perspect Biol. 6:1–19. doi:10.1101/cshperspect.a016428.

Mes-Masson A-M, Witte ON. 1987. Role of The abl Oncogene in Chronic Myelogenous Leukemia. 49. doi: 10.4161/23723548.2014.963450.

Miller RC, Marino SA, Brenner DJ, Martin SG, Richards M, Randers-pehrson G, Hall EJ, Miller RC, Marino SA, Brenner DJ, et al. 1995. The Biological Effectiveness of Radon-Progeny Alpha Particles .142(1):54–60.doi: 10.2307/3578966.

Miller RCM, Randers-pehrson G, Geard CR, Hall EJ, Brenner DJ. 1999. The oncogenic transforming potential of the passage of single a-particles through mammalian cell nuclei. 96(January):19–22.

Minina VI, Soboleva OA, Glushkov AN, Voronina EN, Sokolova EA, Bakanova ML, Savchenko YA, Ryzhkova A V, Titov RA, Druzhinin VG, et al. 2017. and chromosomal aberrations in lung cancer patients. J Cancer Res Clin Oncol. 143(11):2235–2243. doi:10.1007/s00432-017-2486-3.

Mizukami T, Shiraishi K, Shimada Y, Ogiwara H, Tsuta K, Ichikawa H, Sakamoto H, Kato M, Shibata T, Nakano T, et al. 2014. Molecular Mechanisms Underlying Oncogenic RET Fusiont. J Thorac Oncol. 9(5):622–630. doi:10.1097/JTO.0000000000000135.

Monchaux G, Morlier JP, Morin M, Chameaud J. 1994. Carcinogenic and Cocarcinogenic Effects of Radon and Radon Daughters in Rats Masses. Environmental Health Perspectives.102(1):64-73. doi: 10.1289/ehp.9410264.

Moore S, Stanley FKT, Goodarzi AA. 2014. The repair of environmentally relevant DNA double strand breaks caused by high linear energy transfer irradiation – No simple task. DNA repair.
 17:64–73. doi: 10.1016/j.dnarep.2014.01.014.

Morishita M, Muramatsu T, Suto Y, Hirai M. 2016. Chromothripsis-like chromosomal rearrangements induced by ionizing radiation using proton microbeam irradiation system. 7(9). doi:10.18632/oncotarget.7186.

Mukherjee S, Ma Z, Wheeler S, Sathanoori M, Coldren C, Prescott JL, Kozyr N, Bouzyk M, Correll M, Ho H, et al. 2016. Chromosomal microarray provides enhanced targetable gene aberration detection when paired with next generation sequencing panel in profiling lung and colorectal tumors. 209:119–129. doi:10.1016/j.cancergen.2015.12.011.

Muller PAJ, Vousden KH, Norman JC. 2011. p53 and its mutants in tumor cell migration and invasion. 192(2):209–218. doi:10.1083/jcb.201009059.

Murakami H, Keeney S. 2008. Regulating the formation of DNA double-strand breaks in meiosis. Genes Dev. 22(3):286–292. doi:10.1101/gad.1642308.

Nagasawa H, Little JB, Inkret WC, Carpenter S, Thompson K, Raju MR, Chen DJ, Strniste GF. 1990. Cytogenetic effects of extremely low doses of plutonium-238 alpha-particle irradiation in CHO K-1 cells. Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis.244(3):233-8. doi: 10.1016/0165-7992(90)90134-6.

Nagashima H, Shiraishi K, Ohkawa S, Sakamoto Y, Komatsu K, Matsuura S, Tachibana A, Tauchi H. 2018. Induction of somatic mutations by low-dose X-rays : the challenge in recognizing radiation-induced events. 59(October 2017):11–17. doi:10.1093/jrr/rrx053.

Norppa H, Bonassi S, Hansteen I, Hagmar L, Str U, Knudsen LE, Barale R, Fucic A. 2006. Chromosomal aberrations and SCEs as biomarkers of cancer risk. 600:37–45. doi:10.1016/j.mrfmmm.2006.05.030.

NRC. 1990. Health Effects of Exposure to Low Levels of Ionizing Radiation (BEIR V).
Ohshima K, Hatakeyama K, Nagashima T, Watanabe Y, Kanto K, Doi Y, Ide T, Shimoda Y, Tanabe T, Ohnami Sumiko, et al. 2017. Integrated analysis of gene expression and copy number identified potential cancer driver genes with amplification-dependent overexpression in 1,454 solid tumors. Sci Rep. 7(1):641. doi:10.1038/s41598-017-00219-3. [accessed 2019 Jul 25]. http://www.ncbi.nlm.nih.gov/pubmed/28377632.

Okayasu R. 2012. heavy ions — a mini review. 1000:991–1000. doi:10.1002/ijc.26445.
Paik PK, Johnson ML, Angelo SPD, Sima CS, Ang D. 2012. Driver Mutations Determine Survival in Smokers and. doi:10.1002/cncr.27637.

Pal HC, Sharma S, Strickland LR, Agarwal J, Athar M, Elmets A, Afaq F. 2013. Delphinidin Reduces Cell Proliferation and Induces Apoptosis of Non-Small-Cell Lung Cancer Cells by Targeting EGFR / VEGFR2 Signaling Pathways. 8(10):
1–13. doi:10.1371/journal.pone.0077270.

Panov SZ. 2005. Molecular biology of the lung cancer. 39(3):197–210.

Patel KJ, Yu VPCC, Lee H, Corcoran A, Thistlethwaite FC, Evans MJ, Colledge WH, Friedman LS, Ponder BAJ, Venkitaraman AR. 1998. Involvement of Brca2 in DNA Repair. Molecular Cell 1(3):347-57. doi: 10.1016/S1097-2765(00)80035-0.

Paull TT, Rogakou EP, Yamazaki V, Kirchgessner CU, Gellert M, Bonner WM. 2000. A critical role for histone H2AX in recruitment of repair factors to nuclear foci after DNA damage. 10(15):886–895. doi:10.1016/S0960-9822(00)00610-2

Pear BWS, Miller JP, Xu L, Pui JC, Soffer B, Quackenbush RC, Pendergast AM, Bronson R, Aster JC, Scott ML, et al. 1998. Efficient and Rapid Induction of a Chronic Myelogenous Leukemia-Like Myeloproliferative Disease in Mice Receiving P210 bcr/abl-Transduced Bone Marrow. 92(10):3780–3792.

Perera D, Poulos RC, Shah A, Beck D, Pimanda JE, Wong JW. 2016. Differential DNA repair underlies mutation hotspots at. doi:10.1038/nature17437.

Pitot H. 1993. The molecular biology of carcinogenesis. Cancer. 72(S3):962–970. doi:10.1002/1097-0142(19930801)

Polo SE, Jackson SP. 2011. Dynamics of DNA damage response proteins at DNA breaks: a focus on protein modifications. Genes Dev:409–433. doi:10.1101/gad.2021311.c

Povirk LF. 2006. Biochemical mechanisms of chromosomal translocations resulting from DNA double-strand breaks. 5:1199–1212. doi:10.1016/j.dnarep.2006.05.016.

Ptácek O, Stavreva DA, Kim JK, Gichner T. 2001. Induction and repair of DNA damage as measured by the Comet assay and the yield of somatic mutations in gamma-irradiated tobacco seedlings. 491:17–23. doi:10.1016/S1383-5718(00)00146-7.

Pucci B, Kasten M, Giordano A. 2000. Cell Cycle and Apoptosis 1. 2(4):291–299. .doi: 10.1038/sj.neo.7900101.

Ramkissoon A, Navaranjan G, Berriault C, Villeneuve PJ, Demers PA, Do MT. 2018. Histopathologic Analysis of Lung Cancer Incidence Associated with Radon Exposure among Ontario Uranium Miners. doi:10.3390/ijerph15112413.

Robertson A, Allen J, Laney R, Curnow A. 2013. The cellular and molecular carcinogenic effects of radon exposure: A review. International Journal of Molecular Sciences.14(7):14024-63. doi: 10.3390/ijms140714024.

Robertson JB, Koehler A, George J, Little JB. 1983. Oncogenic Transformation of Mouse BALB / 3T3 Cells by Plutonium-238 Alpha Particles. Radiation Research. 96(2):261-74.doi: 10.2307/3576209.

Rode A, Maass KK, Willmund KV, Lichter P. 2016. Chromothripsis in cancer cells : An update. 2333:2322–2333. doi:10.1002/ijc.29888.

Rodríguez-Martínez Á, Torres-Durán M, Barros-Dios JM, Ruano-Ravina A. 2018. Residential radon and small cell lung cancer. A systematic review. Cancer Lett. 426:57–62. doi:10.1016/j.canlet.2018.04.003.
Rogakou EP, Boon C, Redon C, Bonner WM. 1999. Megabase Chromatin Domains Involved in DNA Double-Strand Breaks In Vivo. The Journal of Cell Biology.146(5):905-16. doi: 10.1083/jcb.146.5.905.

Roth JA, Nguyen D, Lawrence DD, Kemp BL, Carrasco CH, Ferson DZ, Hong WK, Romaki R, Lee J., Nesbitt JC, et al. 1996. Retrovirus-mediated wild-type p53 gene transfer to tumors of patients with lung cancer. Nature Medicine.2(9):985-91. doi: 10.1038/nm0996-985.

Rothkamm K, Barnard S, Moquet J, Ellender M, Rana Z, Burdak-rothkamm S. 2015. Review DNA Damage Foci : Meaning and Significance. 504(March). doi:10.1002/em.

Rothkamm K, Lo M. 2003. Evidence for a lack of DNA double-strand break repair in human cells exposed to very low x-ray doses. Proceedings of the National Academy of Sciences.100(9):5057-62. doi: 10.1073/pnas.0830918100.

Rube CE, Grudzenski S, Ku M, Dong X, Rief N, Lo M, Ru C. 2008. Cancer Therapy : Preclinical DNA Double-Strand Break Repair of Blood Lymphocytes and Normal Tissues Analysed in a Preclinical Mouse Model : Implications for Radiosensitivity Testing. 14(20):6546–6556. doi:10.1158/1078-0432.CCR-07-5147.

Russell WL, Conlon MJ, Russell LB, Kelly EM. 1957. Radiation Dose Rate and Mutation Frequency. 128(12). doi: 10.1126/science.128.3338.1546.

Russo A, Pacchierotti F, Cimini D, Ganem NJ, Genesc A, Natarajan AT, Pavanello S, Valle G, Degrassi F. 2015. Review Article Genomic Instability : Crossing Pathways at the Origin of Structural and Numerical Chromosome Changes. 580(March). doi:10.1002/em.

Rydberg B, Cooper B, Cooper PK, Holley WR, Chatterjee A. 2005. Dose-Dependent Misrejoining of Radiation-Induced DNA Double-Strand Breaks in Human Fibroblasts : Experimental and Theoretical Study for High- and Low-LET Radiation. 534:526–534. doi: 10.1667/RR3346.

Sage E, Shikazono N. 2017. Free Radical Biology and Medicine Radiation-induced clustered DNA lesions : Repair and mutagenesis. Free Radic Biol Med. 107(December 2016):125–135. doi:10.1016/j.freeradbiomed.2016.12.008.

San Filippo J, Sung P, Klein H. 2008. Mechanism of Eukaryotic Homologous Recombination. Annu Rev Biochem. 77(1):229–257. doi:10.1146/annurev.biochem.77.061306.125255.

Sanders HR, Albitar M. 2010. Somatic mutations of signaling genes in non-small-cell lung cancer. Cancer Genet Cytogenet. 203(1):7–15. doi:10.1016/j.cancergencyto.2010.07.134.

Santovito A, Cervella P, Delpero M. 2013. Increased frequency of chromosomal aberrations and sister chromatid exchanges in peripheral lymphocytes of radiology technicians chronically exposed to low levels of ionizing radiations. Environ Toxicol Pharmacol. 37(1):396–403. doi:10.1016/j.etap.2013.12.009.

Sasai K, Sukezane T, Yanagita E, Nakagawa H, Hotta A. 2011. Oncogene-Mediated Human Lung Epithelial Cell Transformation Produces Adenocarcinoma Phenotypes In Vivo. (10):2541–2550. doi:10.1158/0008-5472.CAN-10-2221.

Sasaki T, Roding SJ, Chirieac LR, Janne PA. 2010. The Biology and Treatment of EML4-ALK Non-Small Cell Lung Cancer. 46(10):1773–1780. doi:10.1016/j.ejca.2010.04.002.

Sato M, Vaughan MB, Girard L, Peyton M, Lee W, Shames DS, Ramirez RD, Sunaga N, Gazdar AF, Shay JW, et al. 2006. Mutant EGFRs , p16 Bypass , Telomerase ) Are Not Sufficient to Confer a Full Malignant Phenotype on Human Bronchial Epithelial Cells. (4):2116–2129. doi:10.1158/0008-5472.CAN-05-2521.

Sato T, Morita M, Tanaka R, Inoue YUI, Nomura M. 2017. Ex vivo model of non ‑ small cell lung cancer using mouse lung epithelial cells. :6863–6868. doi:10.3892/ol.2017.7098.
Schabath MB, Welsh EA, Fulp WJ, Chen L, Teer JK, Thompson ZJ, Engel BE, Xie M, Berglund AE, Creelan BC, et al. 2016. Differential association of STK11 and TP53 with KRAS. 35(24):3209–3216. doi:10.1038/onc.2015.375.

Schipler A, Iliakis G. 2013. SURVEY AND SUMMARY DNA double-strand – break complexity levels and their possible contributions to the probability for error-prone processing and repair pathway choice. 41(16):7589–7605. doi:10.1093/nar/gkt556.

Schmid E, Regulla D, Kramer H, Harder D. 2002. The Effect of 29 kV X Rays on the Dose Response of Chromosome Aberrations in The Effect of 29 kV X Rays on the Dose Response of Chromosome Aberrations in Human Lymphocytes. 158(6):771–777. doi: 10.1667/0033-7587(2002)158[0771:TEOKXR]2.0.CO;2.

Sheen S, Lee KS, Chung WY, Nam S, Kang DR. 2016. An updated review of case-control studies of lung cancer and indoor radon-Is indoor radon the risk factor for lung cancer? Ann Occup Environ Med. 28(1). doi:10.1186/s40557-016-0094-3.

Shelke S, Das B. 2015. Dose response and adaptive response of non- homologous end joining repair genes and proteins in resting human peripheral blood mononuclear cells exposed to γ radiation. (December 2014):365–379. doi:10.1093/mutage/geu081.

Sherborne AL, Davidson PR, Yu K, Nakamura AO, Rashid M, Nakamura JL, Sherborne AL, Davidson PR, Yu K, Nakamura AO, et al. 2015. Mutational Analysis of Ionizing Radiation Induced Article Mutational Analysis of Ionizing Radiation Induced Neoplasms. CellReports. 12(11):1915–1926. doi:10.1016/j.celrep.2015.08.015.

Shlien A, Malkin D. 2009. Copy number variations and cancer. Genome Medicine.1(6):62.
doi:10.1186/gm62.
Simsek Denis, Jasin M. 2010. HHS Public Access. 118(24):6072–6078. doi:10.1002/cncr.27633.

Simsek Deniz, Jasin M. 2010. Alternative end-joining is suppressed by the canonical NHEJ component Xrcc4/ligase IV during chromosomal translocation formation Deniz. Nat Struct Mol Bio. 17(4):410–416. doi:10.1038/nsmb.

Sishc BJ, Davis AJ. 2017. The Role of the Core Non-Homologous End Joining Factors in Carcinogenesis and Cancer. doi:10.3390/cancers9070081.

Smerhovsky Z, Landa K, Rossner P, Juzova D, Brabec M, Zudova Z, Hola N, Zarska H, Nevsimalova E. 2002. Increased risk of cancer in radon-exposed miners with elevated frequency of chromosomal aberrations. Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis. 514(1-2):165-76. doi: 10.1016/S1383-5718(01)00328-X.

Smith J, Baldeyron C, Oliveira I De, Sala-trepat M, Papadopoulo D. 2001. The influence of DNA double-strand break structure on end-joining in human cells. 29(23):4783–4792. doi: 10.1093/nar/29.23.4783.

Smith J, Riballo E, Kysela B, Baldeyron C, Manolis K, Masson C, Lieber MR, Papadopoulo D, Jeggo P. 2003. Impact of DNA ligase IV on the ® delity of end joining in human cells. 31(8). doi:10.1093/nar/gkg317.

Smith TA, Kirkpatrick DR, Smith S, Smith TK, Pearson T, Kailasam A, Herrmann KZ, Schubertv J, Agrawal DK. 2017. Radioprotective agents to prevent cellular damage due to ionizing radiation. :1–18. doi:10.1186/s12967-017-1338-x.

Soda M, Choi YL, Enomoto M, Takada S, Yamashita Y, Ishikawa S, Fujiwara S, Watanabe H, Kurashina K, Hatanaka H, et al. 2007. Identification of the transforming EML4 – ALK fusion gene in non-small-cell lung cancer. 448(August). doi:10.1038/nature05945.

Somers CM, Sharma R, Quinn JS, Boreham DR. 2004. Gamma radiation-induced heritable mutations at repetitive DNA loci in out-bred mice. 568:69–78. doi:10.1016/j.mrfmmm.2004.06.047.

Stevens ADL, Bradley S, Goodhead DT, Hill MA, Stevens DL, Bradley S, Goodhead T, Hill MA. 2014. The Influence of Dose Rate on the Induction of Chromosome Aberrations and Gene Mutation after Exposure of Plateau Phase V79-4 Cells with High-LET Alpha Particles The Influence of Dose Rate on the Induction of Chromosome Aberrations and Gene Mutation after. 182(3):331–337. doi:10.1667/RR13746.1.

Stopper H, Schmitt E, Gregor C, Mueller SO, Fischer WH. 2003. Increased cell proliferation is associated with genomic instability : elevated micronuclei frequencies in estradiol-treated human ovarian cancer cells hormone ’ s tumor promoting activity . Recently , estradiol are found at higher concentrations. Mutagenesis.18(3):243-7. doi:10.1093/mutage/18.3.243.

Sudprasert W, Navasumrit P, Ruchirawat M. 2006. Effects of low-dose gamma radiation on DNA damage , chromosomal aberration and expression of repair genes in human blood cells. 209:503–511. doi:10.1016/j.ijheh.2006.06.004.

Sun Q, Shi R, Wang X, Li D, Wu H, Ren B. 2016. Biochemical and Biophysical Research Communications Overexpression of ZIC5 promotes proliferation in non-small cell lung cancer. 479:502–509. doi:10.1016/j.bbrc.2016.09.098.

Sutherland BM, Bennett P V, Sidorkina O, Laval J. 2000. Clustered DNA damages induced in isolated DNA and in human cells by low doses of ionizing radiation. Proc Natl Acad Sci U S A. 97(1):103-8. doi: 10.1073/pnas.97.1.103.

Suto Y, Akiyama M, Noda T, Hirai M. 2015. Mutation Research / Genetic Toxicology and Environmental Mutagenesis Construction of a cytogenetic dose – response curve for low-dose range gamma-irradiation in human peripheral blood lymphocytes using. 794:32–38.

Suzuki K, Hei TK. 1996. Mutation induction in gamma-irradiated primary human bronchial epithelial cells and molecular analysis of the HPRT- mutants. 349(I 996). doi: 10.1016/0027-5107(95)00123-9.

Targa A, Rancati G. 2018. ScienceDirect Cancer : a CINful evolution.
Terato H, Ide H. 2005. Clustered DNA damage induced by heavy ion particles. Biol Sci Sp. 18(4):206–215. doi:10.2187/bss.18.206.

Thibervile L, Payne P, Leriche J, Horsman D, Nouvet G, Palcic B, Lam S. 1995. Advances in Brief Evidence of Cumulative Gene Losses with Progression of Premalignant Epithelial Lesions to Carcinoma of the Bronchus. (55):5133–5139.

Thomas P, Umegaki K, Fenech M. 2003. Nucleoplasmic bridges are a sensitive measure of chromosome rearrangement in the cytokinesis-block micronucleus assay Nucleoplasmic bridges are a sensitive measure of chromosome rearrangement in the cytokinesis-block micronucleus assay. (May 2014). doi:10.1093/mutage/18.2.187.

Thompson LL, Jeusset LM, Lepage CC, Mcmanus KJ. 2017. Evolving Therapeutic Strategies to Exploit Chromosome Instability in Cancer. :1–22. doi:10.3390/cancers9110151.

Tirmarchel M, Raphalen A, Allin F, Chameaud J, Bredon P. 1993. Mortality of a cohort of French uranium miners exposed to relatively low radon concentrations. British Journal of Cancer. 67(5):1090-7. doi: 10.1038/bjc.1993.200.

To MD, Quigley DA, Mao J, Rosario R Del, Hsu J, Hodgson G. 2011. Progressive Genomic Instability in the FVB / Kras LA2 Mouse Model of Lung Cancer. :1339–1346. doi:10.1158/1541-7786.MCR-11-0219.

Torres-Durán M, Barros-Dios JM, Fernández-Villar A, Ruano-Ravina A. 2014. Residential radon and lung cancer in never smokers. A systematic review. Cancer Lett. 345(1):21–26. doi:10.1016/j.canlet.2013.12.010.

Trask BJ. 2002. HUMAN CYTOGENETICS : AND COUNTING. 3(October). doi:10.1038/nrg905.
Tu S, Zhang XIALI, Wan HUIF, Xia YANQIN, Liu ZQI. 2018. Effect of taurine on cell proliferation and apoptosis human lung cancer A549 cells. :5473–5480. doi:10.3892/ol.2018.8036.

Tucker JD, Cofield J, Matsumoto K, Ramsey MJ, Freeman DC. 2005a. Persistence of Chromosome Aberrations Following Acute Radiation: I, PAINT Translocations, Dicentrics, Rings, Fragments, and Insertions. 248(January). doi:10.1002/em.20090.

Tucker JD, Cofield J, Matsumoto K, Ramsey MJ, Freeman DC. 2005b. Persistence of Chromosome Aberrations Following Acute Radiation : II , Does It Matter How Translocations Are Scored ? 257(January). doi:10.1002/em.20089.

Vacquier B, Caer S, Rogel A, Feurprier M, Tirmarche M, Luccioni C, Quesne B, Acker A, Laurier D. 2008. Mortality risk in the French cohort of uranium miners : extended follow-up 1946 – 1999. :597–604. doi:10.1136/oem.2007.034959.

Varella-garcia M. 2009. Chromosomal and genomic changes in lung cancer. 4(1):100–106.
Vellingiri B, Shanmugam S, Devi M, Anand S, N DS, Cho S, Keshavarao S. 2014. Ecotoxicology and Environmental Safety Cytogenetic endpoints and Xenobiotic gene polymorphism in lymphocytes of hospital workers chronically exposed to ionizing radiation in Cardiology , Radiology and Orthopedic Laboratories. 100:266–274. doi:10.1016/j.ecoenv.2013.09.036.

Venkitaraman AR. 2002. and the Functions of BRCA1 and BRCA2. 108:171–182.
Vodicka P, Musak L, Vodickova L, Vodenkova S, Vymetalkova V, Försti A, Hemminki K. 2018. Mutat Res Gen Tox En Genetic variation of acquired structural chromosomal aberrations. 836(May):13–21. doi:10.1016/j.mrgentox.2018.05.014.

Ventura A, Kirsch DG, Mclaughlin ME, Tuveson DA, Grimm J, Lintault L, Newman J, Reczek EE, Weissleder R, Jacks T. 2007. Restoration of p53 function leads to tumour regression in vivo. 445(February). doi:10.1038/nature05541.

Vignard J, Mirey G, Salles B. 2013. Ionizing-radiation induced DNA double-strand breaks : A direct and indirect lighting up. 108:362–369. doi:10.1016/j.radonc.2013.06.013.

Vodenkova S, Polivkova Z, Musak L, Smerhovsky Z, Zoubkova H, Sytarova S, Kavcova E, Halasova E, Vodickova L, Jiraskova K, et al. 2015. Structural chromosomal aberrations as potential risk markers in incident cancer patients. (March):557–563.
doi:10.1093/mutage/gev018.

Vodicka P, Musak L, Vodickova L, Vodenkova S, Vymetalkova V, Försti A, Hemminki K. 2018. Mutat Res Gen Tox En Genetic variation of acquired structural chromosomal aberrations. 836(May):13–21. doi:10.1016/j.mrgentox.2018.05.014.

Vogelstein B, Kinzler KW. 2004. OUR TENTH YEAR Cancer genes and the pathways they control. 10(8):789–799. doi:10.1038/nm1087.

Walsh L, Dufey F, Tschense A, Schnelzer M, Grosche B, Kreuzer M. 2010. Radon And The Risk of Cancer Mortality- International Poisson Models For The German Uranium Miners Cohort. (December 2009). doi:10.1097/HP.0b013e3181cd669d.

Wanitchakool P, Jariyawat S, Suksen K, Soorukram D. 2012. Cleistanthoside A tetraacetate-induced DNA damage leading to cell cycle arrest and apoptosis with the involvement of p53 in lung cancer cells. Eur J Pharmacol. 696(1–3):35–42. doi:10.1016/j.ejphar.2012.09.029.

Ward J. F. 1988. DNA Damage Produced by Ionizing Radiation in Mammalian Cells: Identities, Mechanisms of Formation, and Reparability. Prog Nucleic Acid Res Mol Biol. 35(C):95–125. doi:10.1016/S0079-6603(08)60611-X.

Warin RF, Chen H, Soroka DN, Zhu Y, Sang S. 2014. Induction of Lung Cancer Cell Apoptosis through a p53 Pathway by [6]-Shogaol and Its Cysteine-Conjugated Metabolite M2. Journal of Agricultural and Food Chemistry. 62(6). doi:10.1021/jf405573e.

Waters CA, Strande NT, Pryor JM, Strom CN, Mieczkowski P, Burkhalter MD, Oh S, Qaqish BF, Moore DT, Hendrickson EA, et al. 2014. The fidelity of the ligation step determines how ends are resolved during nonhomologous end joining. Nat Commun. 5:1–11. doi:10.1038/ncomms5286.

Weaver DA, Hei TK, Hukku B, DeMuth JP, Crawford EL, McRaven JA, Girgis S, Willey JC. 2000. Localization of tumor suppressor gene candidates by cytogenetic and short tandem repeat analyses in tumorigenic human bronchial epithelial cells Localization of tumor suppressor gene candidates by cytogenetic and short tandem repeat analyses in tumorigeni. (March). doi:10.1093/carcin/21.2.205.

Weaver DA, Hei TK, Hukku B, Mcraven JA, Willey JC. 1997. Cytogenetic and molecular genetic analysis of tumorigenic human bronchial epithelial cells induced by radon alpha particles Cytogenetic and molecular genetic analysis of tumorigenic human bronchial epithelial cells induced by radon alpha particles. Carcinogenesis. 18(6):1251-7

Weinstock DM, Richardson CA, Elliott B, Jasin M. 2006. Modeling oncogenic translocations : Distinct roles for double-strand break repair pathways in translocation formation in mammalian cells. 5:1065–1074. doi:10.1016/j.dnarep.2006.05.028.

Welcker M, Clurman BE. 2008. FBW7 ubiquitin ligase: a tumour suppressor at the crossroads of cell division, growth and differentiation. Nat Publ Gr. 8(february). doi:10.1038/nrc2290.

Werner AE, Wang Y, Doetsch PW. 2017. A Single Exposure to Low- or High-LET Radiation Induces Persistent Genomic Damage in Mouse Epithelial Cells In Vitro and in Lung Tissue A Single Exposure to Low- or High-LET Radiation Induces Persistent Genomic Damage in Mouse Epithelial Cells In Vitro an. 188(4):373–380. doi:10.1667/RR14685.1.

Wessendorf P, Vijg J, Nussenzweig A, Digweed M. 2014. Mutation Research / Fundamental and Molecular Mechanisms of Mutagenesis Deficiency of the DNA repair protein nibrin increases the basal but not the radiation induced mutation frequency in vivo. Mutat Res - Fundam Mol Mech Mutagen. 769:11–16. doi:10.1016/j.mrfmmm.2014.07.001.

Wilhelm T, Magdalou I, Barascu A, Técher H, Debatisse M. 2014. Spontaneous slow replication fork progression elicits mitosis alterations in homologous recombination-deficient mammalian cells. 111(2). doi:10.1073/pnas.1311520111.

Wilson TE, Arlt MF, Park SH, Rajendran S, Paulsen M, Ljungman M, Glover TW. 2015. Large transcription units unify copy number variants and common fragile sites arising under replication stress. :189–200. doi:10.1101/gr.177121.114.Freely.

Winegar RA, Lutze LH, Loughlin KGO, Mirsalis JC. 1994. Radiation-induced point mutations, deletions and micronuclei in lacI transgenic mice. 307:479–487. doi:  10.1016/0027-5107(94)90258-5.

Wistuba II, Behrens C, Milchgrub S, Bryant D, Hung J, Minna JD, Gazdar AF. 1999. Sequential molecular abnormalities are involved in the multistage development of squamous cell lung carcinoma. Oncogene. 18(3):643-50. doi: 10.1038/sj.onc.1202349.

Wrage M, Ruosaari S, Eijk PP, Kaifi JT, Hollme J, Yekebas EF, Izbicki JR, Brakenhoff RH, Streichert T, Riethdorf S, et al. 2009. Human Cancer Biology Genomic Profiles Associated with Early Micrometastasis in Lung Cancer : Relevance of 4q Deletion. 15(5):1566–1575. doi:10.1158/1078-0432.CCR-08-2188.

Yamada AY, Iwata K, Blyth BJ, Doi K, Yamada Y, Iwata K, Blyth BJ, Doi K, Morioka T, Daino K. 2017. Effect of Age at Exposure on the Incidence of Lung and Mammary Cancer after Thoracic X-Ray Irradiation in Wistar Rats Effect of Age at Exposure on the Incidence of Lung and Mammary Cancer after Thoracic X-Ray Irradiation in Wistar Rats. 187(2):210–220. doi:10.1667/RR14478.1.

Yamada Y, Oghiso Y, Enomoto H, Ishigure N. 2002. Induction Of Micronuclei In A Rat Alveolar Epithelia Cell Line By Alpha Particle Irradiation. 99:219–222. doi: 10.1093/oxfordjournals.rpd.a006767.

Zhang N, Wang M, Zhang P, Huang T. 2016. Biochimica et Biophysica Acta Classi fi cation of cancers based on copy number variation landscapes ☆. BBA - Gen Subj. 1860(11):2750–2755. doi:10.1016/j.bbagen.2016.06.003.

Zhong C, Zhou Y, Douglas GC, Witschi H, Ã KEP. 2005. MAPK / AP-1 signal pathway in tobacco smoke-induced cell proliferation and squamous metaplasia in the lungs of rats. 26(12):2187–2195. doi:10.1093/carcin/bgi189.